CCR6 compounds

ABSTRACT

Compounds of formula (I) are provided which are useful in the treatment of diseases or conditions modulated at least in part by CCR6:

CROSS-REFERENCES TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.15/074,965, filed Mar. 18, 2016, which is a continuation of U.S. patentapplication Ser. No. 14/557,949, filed Dec. 2, 2014 (now U.S. Pat. No.9,340,509), which claims the benefit of priority to U.S. PatentApplication No. 61/910,838, filed Dec. 2, 2013, the disclosures of whichare herein incorporated by reference in their entirety.

STATEMENT AS TO RIGHTS TO INVENTIONS MADE UNDER FEDERALLY SPONSOREDRESEARCH AND DEVELOPMENT

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BACKGROUND OF THE INVENTION

Chemokines are chemotactic cytokines that are released by a wide varietyof cells to attract macrophages, T cells, eosinophils, basophils andneutrophils to sites of inflammation (reviewed in Schall, Cytokine,3:165-183 (1991), Schall, et al., Curr Opin. Immunol. 6:865-873 (1994)and Murphy, Rev. Immun., 12:593-633 (1994)). In addition to stimulatingchemotaxis, other changes can be selectively induced by chemokines inresponsive cells, including changes in cell shape, transient rises inthe concentration of intracellular free calcium ions ([Ca2+]), granuleexocytosis, integrin upregulation, formation of bioactive lipids (e.g.,leukotrienes) and respiratory burst, associated with leukocyteactivation. Thus, the chemokines are early triggers of the inflammatoryresponse, causing inflammatory mediator release, chemotaxis andextravasation to sites of infection or inflammation.

There are two main classes of chemokines, CXC (alpha) and CC (beta),depending on whether the first two cysteines are separated by a singleamino acid (C-X-C) or are adjacent (C-C). The alpha-chemokines, such asinterleukin-8 (IL-8), neutrophil-activating protein-2 (NAP-2) andmelanoma growth stimulatory activity protein (MGSA) are chemotacticprimarily for neutrophils, whereas beta-chemokines, such as RANTES,MIP-1a, MIP-1b, monocyte chemotactic protein-1 (MCP-1), MCP-2, MCP-3 andeotaxin are chemotactic for macrophages, T-cells, eosinophils andbasophils (Deng, et al., Nature, 381:661-666 (1996)). The chemokinesbind specific cell-surface receptors belonging to the family ofG-protein-coupled seven-transmembrane-domain proteins (reviewed inHoruk, Trends Pharm. Sci., 15:159-165 (1994)) which are termed“chemokine receptors.”

On binding their cognate ligands, chemokine receptors transduce anintracellular signal though the associated trimeric G protein, resultingin a rapid increase in intracellular calcium concentration. There are atleast eleven human chemokine receptors that bind or respond tobeta-chemokines and at least seven human chemokine receptors that bindto the alpha chemokines. Additionally CX3CR1 (fractalkine receptor) canbind to the fractalkine chemokine, which is distinguished by a series ofthree amino acids between the first two cysteines. Chemokine receptors,have been implicated as being important mediators of inflammatory andimmunoregulatory disorders and diseases, including asthma and allergicdiseases, as well as autoimmune pathologies such as rheumatoid arthritisand atherosclerosis.

CCR6 is known to be expressed primarily in B cells, IL17 secreting Tcells, regulatory T cells and dendritic cells and shows strong bindingto its cognate ligand CCL20 (MIP-3α). It is expressed on approximately30-60% of adult peripheral blood effector/memory CD4+ T cells. CCR6 isinvolved in leukocyte homing to inflamed tissue, particularly the skinand lungs and is co-expressed on almost all T cells that have a skinhoming phenotype, the CLA+ T cells. Thus CCR6 may be an important playerin skin pathologies in which leukocytes participate.

CCR6 expression has been linked to psoriasis in the following manner. Inhumans, a large majority of skin-homing CD4 T cells in the peripheralblood express CCR6 with a greater degree of CCL20-mediated chemotaxisoccurring in T cells isolated from psoriatic patients (Homey, et. al.,JI, 2000). IL17 secreting cells are central agents in severalinflammatory diseases. T cells, such as γδ T cells and TH17 T cellsproduce IL17 after activation. The pathogenic effects of IL17 have beenassociated with human diseases such as rheumatoid arthritis (Patel D Det. al., Ann Rheum Dis 2013), multiple sclerosis (Zepp J, Wu L, and X LiTrends Immunol 2011), and psoriasis (Martin D A et. al., J InvestDermatol 2012). Evidence strongly linking IL17 with psoriasis includegene wide association studies that show strong association betweenpsoriasis and genes upstream (IL-23) or downstream (NFκb) of IL17signaling pathways as well as efficacy in targeting IL17 in a clinicalsetting (Martin D A et. al., J. Invest Dermat. 2012; Papp et. al., NEJM,2012; Papp et. al., NEJM, 2012). In addition to enhanced CCL20-mediatedchemotaxis, CCR6+ T cells isolated from psoriatic patientspreferentially secrete IL-17A, IL22, and TNFα when compared to healthycontrols (Kagami, et. al., J. Invest. Dermatol., 2010). Lastly, ccl20mRNA was up-regulated in lesional psoriatic skin samples (Homey, et.al., JI, 2000; Dieu-Nosjean, et. al., JEM, 2000). In mice, CCR6knock-out mice were protected from IL-23 driven psoriasis. Thus, amultitude of evidence in both mice and men suggest a protective role forCCR6 blockade in psoriasis and psoriasis-like models.

In view of the clinical importance of CCR6, the identification ofcompounds that modulate CCR6 function represent an attractive avenueinto the development of new therapeutic agents. Such compounds andmethods for their use are provided herein.

BRIEF SUMMARY OF THE INVENTION

Described herein are compounds having formula (I):

wherein the ring vertices a, b, c and d; Ar; A; R², R³ and thesubscripts m and n have the meanings provided in the DetailedDescription below. The compounds have utility in the treatment ofdiseases or conditions modulated at least in part by CCR6.

Pharmaceutical compositions of the compounds of formula (I) are alsoprovided.

Further provided in the present disclosure preparative methods for thesynthesis of compounds of formula (I), as well as selected intermediatesuseful in the preparation.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGS. 1A-1H provide structures and binding data for compounds preparedand evaluated by the methods described herein. Activity is shown as +,20000 nM≥IC₅₀≥500 nM; ++, 500 nM>IC₅₀≥100 nM; +++, 100 nM>IC₅₀.

FIGS. 2A-2C provide structures and migration data for compounds preparedand evaluated by the methods described herein. Activity is shown as +,20000 nM≥IC50≥500 nM; ++, 500 nM>IC50≥100 nM; +++, 100 nM>IC50.

DETAILED DESCRIPTION OF THE INVENTION I. Abbreviation and Definitions

The term “alkyl”, by itself or as part of another substituent, means,unless otherwise stated, a straight or branched chain hydrocarbonradical, having the number of carbon atoms designated (i.e. C₁₋₈ meansone to eight carbons). Examples of alkyl groups include methyl, ethyl,n-propyl, isopropyl, n-butyl, t-butyl, isobutyl, sec-butyl, n-pentyl,n-hexyl, n-heptyl, n-octyl, and the like. The term “alkenyl” refers toan unsaturated alkyl group having one or more double bonds. Similarly,the term “alkynyl” refers to an unsaturated alkyl group having one ormore triple bonds. Examples of such unsaturated alkyl groups includevinyl, 2-propenyl, crotyl, 2-isopentenyl, 2-(butadienyl),2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1- and 3-propynyl,3-butynyl, and the higher homologs and isomers. The term “cycloalkyl”refers to hydrocarbon rings having the indicated number of ring atoms(e.g., C₃₋₆cycloalkyl) and being fully saturated or having no more thanone double bond between ring vertices. “Cycloalkyl” is also meant torefer to bicyclic and polycyclic hydrocarbon rings such as, for example,bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, etc. The term“heterocycloalkane” or “heterocycloalkyl” refers to a cycloalkyl groupthat contain from one to five heteroatoms selected from N, O, and S,wherein the nitrogen and sulfur atoms are optionally oxidized, and thenitrogen atom(s) are optionally quaternized. The heterocycloalkane maybe a monocyclic, a bicyclic or a polycylic ring system. Non limitingexamples of heterocycloalkane groups include pyrrolidine, imidazolidine,pyrazolidine, butyrolactam, valerolactam, imidazolidinone, hydantoin,dioxolane, phthalimide, piperidine, 1,4-dioxane, morpholine,thiomorpholine, thiomorpholine-S-oxide, thiomorpholine-S,S-oxide,piperazine, pyran, pyridone, 3-pyrroline, thiopyran, pyrone,tetrahydrofuran, tetrhydrothiophene, quinuclidine, and the like. Aheterocycloalkane group can be attached to the remainder of the moleculethrough a ring carbon or a heteroatom.

The term “alkylene” by itself or as part of another substituent means adivalent radical derived from an alkane, as exemplified by—CH₂CH₂CH₂CH₂—. Typically, an alkyl (or alkylene) group will have from 1to 24 carbon atoms, with those groups having 10 or fewer carbon atomsbeing preferred in the present invention. A “lower alkyl” or “loweralkylene” is a shorter chain alkyl or alkylene group, generally havingfour or fewer carbon atoms. Similarly, “alkenylene” and “alkynylene”refer to the unsaturated forms of “alkylene” having double or triplebonds, respectively.

As used herein, a wavy line, “

”, that intersects a single, double or triple bond in any chemicalstructure depicted herein, represent the point attachment of the single,double, or triple bond to the remainder of the molecule.

The terms “alkoxy,” “alkylamino” and “alkylthio” (or thioalkoxy) areused in their conventional sense, and refer to those alkyl groupsattached to the remainder of the molecule via an oxygen atom, an aminogroup, or a sulfur atom, respectively. Additionally, for dialkylaminogroups, the alkyl portions can be the same or different and can also becombined to form a 3-7 membered ring with the nitrogen atom to whicheach is attached. Accordingly, a group represented as dialkylamino or—NR^(a)R^(b) is meant to include piperidinyl, pyrrolidinyl, morpholinyl,azetidinyl and the like.

The term “di-(C₁₋₄ alkyl)amino-C₁₋₄ alkyl” refers to an amino groupbearing two C₁₋₄ alkyl groups that can be the same or different (e.g.,methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl, isobutyl andtert-butyl) and which is attached to the remainder of the moleculethrough a C₁₋₄ alkyl group (a one to four carbon alkylene linkinggroup). Examples of di-(C₁₋₄ alkyl)amino-C₁₋₄ alkyl groups includedimethylaminomethyl, 2-(ethyl(methyl)amino)ethyl,3-(dimethylamino)butyl, and the like.

The terms “halo” or “halogen,” by themselves or as part of anothersubstituent, mean, unless otherwise stated, a fluorine, chlorine,bromine, or iodine atom. Additionally, terms such as “haloalkyl,” aremeant to include monohaloalkyl and polyhaloalkyl. For example, the term“C₁₋₄ haloalkyl” is mean to include trifluoromethyl,2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like.

The term “aryl” means, unless otherwise stated, a polyunsaturated,typically aromatic, hydrocarbon group which can be a single ring ormultiple rings (up to three rings) which are fused together or linkedcovalently. The term “heteroaryl” refers to aryl groups (or rings) thatcontain from one to five heteroatoms selected from N, O, and S, whereinthe nitrogen and sulfur atoms are optionally oxidized, and the nitrogenatom(s) are optionally quaternized. A heteroaryl group can be attachedto the remainder of the molecule through a heteroatom. Non-limitingexamples of aryl groups include phenyl, naphthyl and biphenyl, whilenon-limiting examples of heteroaryl groups include pyridyl, pyridazinyl,pyrazinyl, pyrimindinyl, triazinyl, quinolinyl, quinoxalinyl,quinazolinyl, cinnolinyl, phthalazinyl, benzotriazinyl, purinyl,benzimidazolyl, benzopyrazolyl, benzotriazolyl, benzisoxazolyl,isobenzofuryl, isoindolyl, indolizinyl, benzotriazinyl, thienopyridinyl,thienopyrimidinyl, pyrazolopyrimidinyl, imidazopyridines,benzothiaxolyl, benzofuranyl, benzothienyl, indolyl, quinolyl,isoquinolyl, isothiazolyl, pyrazolyl, indazolyl, pteridinyl, imidazolyl,triazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiadiazolyl, pyrrolyl,thiazolyl, furyl, thienyl and the like. Substituents for each of theabove noted aryl and heteroaryl ring systems are selected from the groupof acceptable substituents described below.

The term “arylalkyl” is meant to include those radicals in which an arylgroup is attached to an alkyl group (e.g., benzyl, phenethyl, and thelike). Similarly, the term “heteroaryl-alkyl”is meant to include thoseradicals in which a heteroaryl group is attached to an alkyl group(e.g., pyridylmethyl, thiazolylethyl, and the like).

As used herein, the term “heteroatom” is meant to include oxygen (O),nitrogen (N), sulfur (S) and silicon (Si).

The term “pharmaceutically acceptable salts” is meant to include saltsof the active compounds which are prepared with relatively nontoxicacids or bases, depending on the particular substituents found on thecompounds described herein. When compounds of the present inventioncontain relatively acidic functionalities, base addition salts can beobtained by contacting the neutral form of such compounds with asufficient amount of the desired base, either neat or in a suitableinert solvent. Examples of salts derived frompharmaceutically-acceptable inorganic bases include aluminum, ammonium,calcium, copper, ferric, ferrous, lithium, magnesium, manganic,manganous, potassium, sodium, zinc and the like. Salts derived frompharmaceutically-acceptable organic bases include salts of primary,secondary and tertiary amines, including substituted amines, cyclicamines, naturally-occurring amines and the like, such as arginine,betaine, caffeine, choline, N,N′-dibenzylethylenediamine, diethylamine,2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine,ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine,glucosamine, histidine, hydrabamine, isopropylamine, lysine,methylglucamine, morpholine, piperazine, piperidine, polyamine resins,procaine, purines, theobromine, triethylamine, trimethylamine,tripropylamine, tromethamine and the like. When compounds of the presentinvention contain relatively basic functionalities, acid addition saltscan be obtained by contacting the neutral form of such compounds with asufficient amount of the desired acid, either neat or in a suitableinert solvent. Examples of pharmaceutically acceptable acid additionsalts include those derived from inorganic acids like hydrochloric,hydrobromic, nitric, carbonic, monohydrogencarbonic, phosphoric,monohydrogenphosphoric, dihydrogenphosphoric, sulfuric,monohydrogensulfuric, hydriodic, or phosphorous acids and the like, aswell as the salts derived from relatively nontoxic organic acids likeacetic, propionic, isobutyric, malonic, benzoic, succinic, suberic,fumaric, mandelic, phthalic, benzenesulfonic, p-tolylsulfonic, citric,tartaric, methanesulfonic, and the like. Also included are salts ofamino acids such as arginate and the like, and salts of organic acidslike glucuronic or galactunoric acids and the like (see, for example,Berge, S. M., et al, “Pharmaceutical Salts”, Journal of PharmaceuticalScience, 1977, 66, 1-19). Certain specific compounds of the presentinvention contain both basic and acidic functionalities that allow thecompounds to be converted into either base or acid addition salts.

The neutral forms of the compounds may be regenerated by contacting thesalt with a base or acid and isolating the parent compound in theconventional manner. The parent form of the compound differs from thevarious salt forms in certain physical properties, such as solubility inpolar solvents, but otherwise the salts are equivalent to the parentform of the compound for the purposes of the present invention.

In addition to salt forms, the present invention provides compoundswhich are in a prodrug form. Prodrugs of the compounds described hereinare those compounds that readily undergo chemical changes underphysiological conditions to provide the compounds of the presentinvention. Additionally, prodrugs can be converted to the compounds ofthe present invention by chemical or biochemical methods in an ex vivoenvironment. For example, prodrugs can be slowly converted to thecompounds of the present invention when placed in a transdermal patchreservoir with a suitable enzyme or chemical reagent.

Certain compounds of the present invention can exist in unsolvated formsas well as solvated forms, including hydrated forms. In general, thesolvated forms are equivalent to unsolvated forms and are intended to beencompassed within the scope of the present invention. Certain compoundsof the present invention may exist in multiple crystalline or amorphousforms. In general, all physical forms are equivalent for the usescontemplated by the present invention and are intended to be within thescope of the present invention.

Certain compounds of the present invention possess asymmetric carbonatoms (optical centers) or double bonds, the racemates, diastereomers,geometric isomers, regioisomers and individual isomers (e.g., separateenantiomers) are all intended to be encompassed within the scope of thepresent invention. The compounds of the present invention may alsocontain unnatural proportions of atomic isotopes at one or more of theatoms that constitute such compounds. Unnatural proportions of anisotope may be defined as ranging from the amount found in nature to anamount consisting of 100% of the atom in question. For example, thecompounds may incorporate radioactive isotopes, such as for exampletritium (³H), iodine-125 (¹²⁵I) or carbon-14 (¹⁴C), or non-radioactiveisotopes, such as deuterium (²H) or carbon-13 (¹³C). Such isotopicvariations can provide additional utilities to those described elsewherewith this application. For instance, isotopic variants of the compoundsof the invention may find additional utility, including but not limitedto, as diagnostic and/or imaging reagents, or as cytotoxic/radiotoxictherapeutic agents. Additionally, isotopic variants of the compounds ofthe invention can have altered pharmacokinetic and pharmacodynamiccharacteristics which can contribute to enhanced safety, tolerability orefficacy during treatment. All isotopic variations of the compounds ofthe present invention, whether radioactive or not, are intended to beencompassed within the scope of the present invention.

The term “and acid isosteres” means, unless otherwise stated, a groupwhich can replace a carboxylic acid, having an acidic functionality andsteric and electronic characteristics that provide a level of activity(or other compound characteristic such as solubility) similar to acarboxylic acid. Representative acid isosteres include, hydroxamicacids, sulfonic acids, sulfinic acids, sulfonamides, acyl-sulfonamides,phosphonic acids, phosphinic acids, phosphoric acids, tetrazole, andoxo-oxadiazoles.

Compounds of the invention having formula I can exist in differentisomeric forms. As used herein, the terms cis or trans are used in theirconventional sense in the chemical arts, i.e., referring to the positionof the substituents to one another relative to a reference plane, e.g.,a double bond, or a ring system, such as a decalin-type ring system or ahydroquinolone ring system: in the cis isomer, the substituents are onthe same side of the reference plane, in the trans isomer thesubstituents are on opposite sides. Additionally, different conformersare contemplated by the present invention, as well as distinct rotamers.Conformers are conformational isomers that can differ by rotations aboutone or more a bonds. Rotamers are conformers that differ by rotationabout only a single a bond.

II. General

The present invention derives from the discovery that compounds offormula I act as potent antagonists of the CCR6 receptor. The compoundshave in vivo anti-inflammatory activity and have superiorpharmacokinetic properties. Accordingly, the compounds provided hereinare useful in pharmaceutical compositions, methods for the treatment ofCCR6-mediated diseases, and as controls in assays for the identificationof competitive CCR6 antagonists.

III. Compounds

In one aspect, the present invention provides for a compound of FormulaI:

or a pharmaceutically acceptable salt, hydrate, solvate, N-oxide orrotamer thereof. In Formula (I), the letter A represents a carboxylicacid moiety or a carboxylic acid isostere; ring vertices a, b, c and dare independently selected from the group consisting of N, CH and C(R¹);each R¹ is independently selected from the group consisting of halogen,CN, —SF₅, C₁₋₈ alkyl, C₃₋₈ cycloalkyl, C₂₋₈ alkenyl, C₂₋₈ alkynyl, C₁₋₈haloalkyl, —OR^(a), —SR^(a), —COR^(a), —NR^(a)R^(b), and 5- or6-membered heteroaryl, wherein the alkyl portions of R¹ are optionallyfurther substituted with 1-3 R^(a); and optionally, adjacent R¹ membersare connected to form an additional 5- or 6-membered ring which issaturated or unsaturated having ring vertices selected from C, O, S andN, wherein the additional 5- or 6-membered ring is optionallysubstituted with one or two members selected from the group consistingof halogen, hydroxyl, C₁₋₄ alkyl, C₁₋₄ alkoxy and C₁₋₄ haloalkyl; thesubscript m is an integer of from 0 to 2; the subscript n is an integerof from 0 to 3; each R² and R³ is independently selected from the groupconsisting of halogen, CN, C₁₋₈ alkyl, C₃₋₈ cycloalkyl, C₂₋₈ alkenyl,C₂₋₈ alkynyl, C₁₋₈ haloalkyl, aryl, —OR^(a), —NR^(a)R^(b) and—N(R^(a))—C₁-C₄ alkylene-OR^(b), and wherein the alkyl or aryl portionsof R² and R³ are optionally further substituted with 1-3 R^(a); Ar is a5- or 6-membered aromatic or heteroaromatic ring that is optionallysubstituted with from 1 to 5 R⁴ substitutents independently selectedfrom the group consisting of halogen, CN, —SF₅, C₁₋₈ alkyl, C₃₋₈cycloalkyl, C₂₋₈ alkenyl, C₂₋₈ alkynyl, C₁₋₈ haloalkyl, C₁₋₈hydroxyalkyl, —OR³, —NR^(a)R^(b), 5- or 6-membered heteroaryl, and 3-,4-, 5- or 6-membered heterocycloalkane wherein the heteroatoms presentas ring vertices of the heteroaryl and heterocycloalkane rings areselected from N, O and S, and wherein the alkyl, cycloalkyl, aryl,heteroaryl and hetereocycloalkane portions of R⁴ are optionally furthersubstituted with 1-3 R^(a); each R^(a) and R^(b) is independentlyselected from the group consisting of hydrogen, hydroxyl, halogen,cyano, C₁₋₄ alkyl, C₁₋₄ alkoxy, C₁₋₄ haloalkyl, C₃₋₆ cycloalkyl, amino,C₁₋₈ alkylamino, and di C₁₋₈ alkylamino, or when attached to a nitrogenatom are optionally combined to form a 4- to 7-membered saturated ring,which is optionally substituted with oxo.

In some selected embodiments, the compounds of formula (I) are thosewherein ring vertices a, b, c and d are each independently selected fromthe group consisting of CH and C(R¹). In other selected embodiments, thecompounds of formula (I) are those wherein ring vertex d is CH. In stillother selected embodiments, the compounds of formula (I) are thosewherein ring vertices c and d are each CH. In yet other selectedembodiments, the compounds of formula (I) are those wherein ringvertices a, b, and c are each C(R¹). In other selected embodiments, thecompounds of formula (I) are those wherein ring vertices a and b areeach C(R¹).

In the compounds of formula (I), the letter A represents a carboxylicacid or a carboxylic acid isostere (or a suitable salt thereof). Incertain selected embodiments of formula (I), the letter A represents amember selected from the group consisting of —CO₂H, —SO₃H, —PO₃H₂ and atetrazole. In other selected embodiments, the compounds of formula (I)are those wherein A is a carboxylic acid (—CO₂H) or a tetrazole. In someembodiments A is a tetrazole. In some embodiments, A is —CO₂H or a saltthereof.

Turning next to the substituents on the quinoline moiety, in certainselected embodiments m and n are both 0. In other selected embodiments,the compounds of formula (I) are those wherein m and n are independentlyselected from 0 and 1. For those embodiments wherein R² and R³ arepresent, selected embodiments are those wherein each is independentlyselected from the group consisting of halogen, C₁₋₈ alkyl, —OR^(a),—NR^(a)R^(b) and —N(R^(a))—C₁-C₄ alkylene-OR^(b), and wherein the alkylportions of R² and R³ are optionally further substituted with 1-3 R^(a).

In other selected embodiments for the compounds of formula (I), and foreach of noted embodiments above, Ar is a 6-membered aromatic orheteroaromatic ring selected from the group consisting of benzene,pyridine and pyrimidine, each of which is optionally substituted withfrom 1-3 R⁴ substituents. In still other selected embodiments for thecompounds of formula (I), and for each of noted embodiments above, Ar isselected from the group consisting of benzene and pyridine, each ofwhich is optionally substituted with from 1-2 R⁴ substituents.

In one selected group of embodiments of formula (I), Ar is a benzenering which is optionally substituted with from 1-3 R⁴ substituents; A is—CO₂H; d is CH; and each of a, b and c is independently selected fromthe group consisting of CH and C(R¹). In this group of embodiments,further selected embodiments are those wherein m and n are eachindependently 0 or 1. Still further selected embodiments are thosewherein m and n are each 0.

In other selected embodiments for the compounds of formula (I), and foreach of noted embodiments above, the ring having a, b, c and d as ringvertices is selected from the group consisting of:

wherein the wavy line labeled y indicates the point of attachment to theNHS(O)₂ portion of the compound and the wavy line labeled z indicatesthe point of attachment to the quinoline ring.

In other selected embodiments for the compounds of formula (I), and foreach of noted embodiments above, Ar is selected from the groupconsisting of:

wherein the wavy line labeled w indicates the point of attachment to theS(O)₂ moiety.

In still other selected embodiments, the compounds of formula (I) areselected from the group consisting of:

In still other selected embodiments, the compounds of formula (I) areselected from the group consisting of:

or a pharmaceutically acceptable salt, hydrate, solvate, N-oxide orrotamer thereof.Preparation of Compounds

The schemes in the Examples below provide certain synthetic routes thatcan be followed to access certain compounds of the present invention.Other routes or modification of the routes presented below would bereadily apparent to a skilled artisan and are within the scope of thepresent invention.

IV. Pharmaceutical Compositions

In addition the compounds provided above, the compositions formodulating CCR6, activity in humans and animals will typically contain apharmaceutical carrier or diluent.

The term “composition” as used herein is intended to encompass a productcomprising the specified ingredients in the specified amounts, as wellas any product which results, directly or indirectly, from combinationof the specified ingredients in the specified amounts. By“pharmaceutically acceptable” it is meant the carrier, diluent orexcipient must be compatible with the other ingredients of theformulation and not deleterious to the recipient thereof.

The pharmaceutical compositions for the administration of the compoundsof this invention may conveniently be presented in unit dosage form andmay be prepared by any of the methods well known in the art of pharmacyand drug delivery. All methods include the step of bringing the activeingredient into association with the carrier which constitutes one ormore accessory ingredients. In general, the pharmaceutical compositionsare prepared by uniformly and intimately bringing the active ingredientinto association with a liquid carrier or a finely divided solid carrieror both, and then, if necessary, shaping the product into the desiredformulation. In the pharmaceutical composition the active objectcompound is included in an amount sufficient to produce the desiredeffect upon the process or condition of diseases.

The pharmaceutical compositions containing the active ingredient may bein a form suitable for oral use, for example, as tablets, troches,lozenges, aqueous or oily suspensions, dispersible powders or granules,emulsions and self emulsifications as described in U.S. Pat. No.6,451,339, hard or soft capsules, syrups, elixirs, solutions, buccalpatch, oral gel, chewing gum, chewable tablets, effervescent powder andeffervescent tablets. Compositions intended for oral use may be preparedaccording to any method known to the art for the manufacture ofpharmaceutical compositions and such compositions may contain one ormore agents selected from the group consisting of sweetening agents,flavoring agents, coloring agents, antioxidants and preserving agents inorder to provide pharmaceutically elegant and palatable preparations.Tablets contain the active ingredient in admixture with non-toxicpharmaceutically acceptable excipients which are suitable for themanufacture of tablets. These excipients may be for example, inertdiluents, such as cellulose, silicon dioxide, aluminum oxide, calciumcarbonate, sodium carbonate, glucose, mannitol, sorbitol, lactose,calcium phosphate or sodium phosphate; granulating and disintegratingagents, for example, corn starch, or alginic acid; binding agents, forexample PVP, cellulose, PEG, starch, gelatin or acacia, and lubricatingagents, for example magnesium stearate, stearic acid or talc. Thetablets may be uncoated or they may be coated, enterically or otherwise,by known techniques to delay disintegration and absorption in thegastrointestinal tract and thereby provide a sustained action over alonger period. For example, a time delay material such as glycerylmonostearate or glyceryl distearate may be employed. They may also becoated by the techniques described in the U.S. Pat. Nos. 4,256,108;4,166,452; and U.S. Pat. No. 4,265,874 to form osmotic therapeutictablets for control release.

Formulations for oral use may also be presented as hard gelatin capsuleswherein the active ingredient is mixed with an inert solid diluent, forexample, calcium carbonate, calcium phosphate or kaolin, or as softgelatin capsules wherein the active ingredient is mixed with water or anoil medium, for example peanut oil, liquid paraffin, or olive oil.Additionally, emulsions can be prepared with a non-water miscibleingredient such as oils and stabilized with surfactants such asmono-diglycerides, PEG esters and the like.

Aqueous suspensions contain the active materials in admixture withexcipients suitable for the manufacture of aqueous suspensions. Suchexcipients are suspending agents, for example sodiumcarboxymethylcellulose, methylcellulose, hydroxy-propylmethylcellulose,sodium alginate, polyvinyl-pyrrolidone, gum tragacanth and gum acacia;dispersing or wetting agents may be a naturally-occurring phosphatide,for example lecithin, or condensation products of an alkylene oxide withfatty acids, for example polyoxy-ethylene stearate, or condensationproducts of ethylene oxide with long chain aliphatic alcohols, forexample heptadecaethyleneoxycetanol, or condensation products ofethylene oxide with partial esters derived from fatty acids and ahexitol such as polyoxyethylene sorbitol monooleate, or condensationproducts of ethylene oxide with partial esters derived from fatty acidsand hexitol anhydrides, for example polyethylene sorbitan monooleate.The aqueous suspensions may also contain one or more preservatives, forexample ethyl, or n-propyl, p-hydroxybenzoate, one or more coloringagents, one or more flavoring agents, and one or more sweetening agents,such as sucrose or saccharin.

Oily suspensions may be formulated by suspending the active ingredientin a vegetable oil, for example arachis oil, olive oil, sesame oil orcoconut oil, or in a mineral oil such as liquid paraffin. The oilysuspensions may contain a thickening agent, for example beeswax, hardparaffin or cetyl alcohol. Sweetening agents such as those set forthabove, and flavoring agents may be added to provide a palatable oralpreparation. These compositions may be preserved by the addition of ananti-oxidant such as ascorbic acid.

Dispersible powders and granules suitable for preparation of an aqueoussuspension by the addition of water provide the active ingredient inadmixture with a dispersing or wetting agent, suspending agent and oneor more preservatives. Suitable dispersing or wetting agents andsuspending agents are exemplified by those already mentioned above.Additional excipients, for example sweetening, flavoring and coloringagents, may also be present.

The pharmaceutical compositions of the invention may also be in the formof oil-in-water emulsions. The oily phase may be a vegetable oil, forexample olive oil or arachis oil, or a mineral oil, for example liquidparaffin or mixtures of these. Suitable emulsifying agents may benaturally-occurring gums, for example gum acacia or gum tragacanth,naturally-occurring phosphatides, for example soy bean, lecithin, andesters or partial esters derived from fatty acids and hexitolanhydrides, for example sorbitan monooleate, and condensation productsof the said partial esters with ethylene oxide, for examplepolyoxyethylene sorbitan monooleate. The emulsions may also containsweetening and flavoring agents.

Syrups and elixirs may be formulated with sweetening agents, for exampleglycerol, propylene glycol, sorbitol or sucrose. Such formulations mayalso contain a demulcent, a preservative and flavoring and coloringagents. Oral solutions can be prepared in combination with, for example,cyclodextrin, PEG and surfactants.

The pharmaceutical compositions may be in the form of a sterileinjectable aqueous or oleagenous suspension. This suspension may beformulated according to the known art using those suitable dispersing orwetting agents and suspending agents which have been mentioned above.The sterile injectable preparation may also be a sterile injectablesolution or suspension in a non-toxic parenterally-acceptable diluent orsolvent, for example as a solution in 1,3-butane diol. Among theacceptable vehicles and solvents that may be employed are water,Ringer's solution and isotonic sodium chloride solution. In addition,sterile, fixed oils are conventionally employed as a solvent orsuspending medium. For this purpose any bland fixed oil may be employedincluding synthetic mono- or diglycerides. In addition, fatty acids suchas oleic acid find use in the preparation of injectables.

The compounds of the present invention may also be administered in theform of suppositories for rectal administration of the drug. Thesecompositions can be prepared by mixing the drug with a suitablenon-irritating excipient which is solid at ordinary temperatures butliquid at the rectal temperature and will therefore melt in the rectumto release the drug. Such materials include cocoa butter andpolyethylene glycols. Additionally, the compounds can be administeredvia ocular delivery by means of solutions or ointments. Still further,transdermal delivery of the subject compounds can be accomplished bymeans of iontophoretic patches and the like. For topical use, creams,ointments, jellies, solutions or suspensions, etc., containing thecompounds of the present invention are employed. As used herein, topicalapplication is also meant to include the use of mouth washes andgargles.

The compounds of the invention may be formulated for depositing into amedical device, which may include any of variety of conventional grafts,stents, including stent grafts, catheters, balloons, baskets or otherdevice that can be deployed or permanently implanted within a bodylumen. As a particular example, it would be desirable to have devicesand methods which can deliver compounds of the invention to the regionof a body which has been treated by interventional technique.

In exemplary embodiment, the inhibitory agent of this invention may bedeposited within a medical device, such as a stent, and delivered to thetreatment site for treatment of a portion of the body.

Stents have been used as delivery vehicles for therapeutic agents (i.e.,drugs). Intravascular stents are generally permanently implanted incoronary or peripheral vessels. Stent designs include those of U.S. Pat.No. 4,733,655 (Palmaz), U.S. Pat. No. 4,800,882 (Gianturco), or U.S.Pat. No. 4,886,062 (Wiktor). Such designs include both metal andpolymeric stents, as well as self-expanding and balloon-expandablestents. Stents may also used to deliver a drug at the site of contactwith the vasculature, as disclosed in U.S. Pat. No. 5,102,417 (Palmaz)and in International Patent Application Nos. WO 91/12779 (Medtronic,Inc.) and WO 90/13332 (Cedars-Sanai Medical Center), U.S. Pat. No.5,419,760 (Narciso, Jr.) and U.S. Pat. No. 5,429,634 (Narciso, Jr.), forexample. Stents have also been used to deliver viruses to the wall of alumen for gene delivery, as disclosed in U.S. Pat. No. 5,833,651(Donovan et al.).

The term “deposited” means that the inhibitory agent is coated,adsorbed, placed, or otherwise incorporated into the device by methodsknown in the art. For example, the inhibitory agent may be embedded andreleased from within (“matrix type”) or surrounded by and releasedthrough (“reservoir type”) polymer materials that coat or span themedical device. In the later example, the inhibitory agent may beentrapped within the polymer materials or coupled to the polymermaterials using one or more the techniques for generating such materialsknown in the art. In other formulations, the inhibitory agent may belinked to the surface of the medical device without the need for acoating by means of detachable bonds and release with time, can beremoved by active mechanical or chemical processes, or are in apermanently immobilized form that presents the inhibitory agent at theimplantation site.

In one embodiment, the inhibitory agent may be incorporated with polymercompositions during the formation of biocompatible coatings for medicaldevices, such as stents. The coatings produced from these components aretypically homogeneous and are useful for coating a number of devicesdesigned for implantation.

The polymer may be either a biostable or a bioabsorbable polymerdepending on the desired rate of release or the desired degree ofpolymer stability, but a bioabsorbable polymer is preferred for thisembodiment since, unlike a biostable polymer, it will not be presentlong after implantation to cause any adverse, chronic local response.Bioabsorbable polymers that could be used include, but are not limitedto, poly(L-lactic acid), polycaprolactone, polyglycolide (PGA),poly(lactide-co-glycolide) (PLLA/PGA), poly(hydroxybutyrate),poly(hydroxybutyrate-co-valerate), polydioxanone, polyorthoester,polyanhydride, poly(glycolic acid), poly(D-lactic acid), poly(L-lacticacid), poly(D,L-lactic acid), poly(D,L-lactide) (PLA), poly (L-lactide)(PLLA), poly(glycolic acid-co-trimethylene carbonate) (PGA/PTMC),polyethylene oxide (PEO), polydioxanone (PDS), polyphosphoester,polyphosphoester urethane, poly(amino acids), cyanoacrylates,poly(trimethylene carbonate), poly(iminocarbonate), copoly(ether-esters)(e.g., PEO/PLA), polyalkylene oxalates, polyphosphazenes andbiomolecules such as fibrin, fibrinogen, cellulose, starch, collagen andhyaluronic acid, polyepsilon caprolactone, polyhydroxy butyric acid,polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates,cross linked or amphipathic block copolymers of hydrogels, and othersuitable bioabsorbable poplymers known in the art. Also, biostablepolymers with a relatively low chronic tissue response such aspolyurethanes, silicones, and polyesters could be used and otherpolymers could also be used if they can be dissolved and cured orpolymerized on the medical device such as polyolefins, polyisobutyleneand ethylene-alphaolefin copolymers; acrylic polymers and copolymers,vinyl halide polymers and copolymers, such as polyvinyl chloride;polyvinylpyrrolidone; polyvinyl ethers, such as polyvinyl methyl ether;polyvinylidene halides, such as polyvinylidene fluoride andpolyvinylidene chloride; polyacrylonitrile, polyvinyl ketones; polyvinylaromatics, such as polystyrene, polyvinyl esters, such as polyvinylacetate; copolymers of vinyl monomers with each other and olefins, suchas ethylene-methyl methacrylate copolymers, acrylonitrile-styrenecopolymers, ABS resins, and ethylene-vinyl acetate copolymers; pyrancopolymer; polyhydroxy-propyl-methacrylamide-phenol;polyhydroxyethyl-aspartamide-phenol; polyethyleneoxide-polylysinesubstituted with palmitoyl residues; polyamides, such as Nylon 66 andpolycaprolactam; alkyd resins, polycarbonates; polyoxymethylenes;polyimides; polyethers; epoxy resins, polyurethanes; rayon;rayon-triacetate; cellulose, cellulose acetate, cellulose butyrate;cellulose acetate butyrate; cellophane; cellulose nitrate; cellulosepropionate; cellulose ethers; and carboxymethyl cellulose.

Polymers and semipermeable polymer matrices may be formed into shapedarticles, such as valves, stents, tubing, prostheses and the like.

In one embodiment of the invention, the inhibitory agent of theinvention is coupled to a polymer or semipermeable polymer matrix thatis formed as a stent or stent-graft device.

Typically, polymers are applied to the surface of an implantable deviceby spin coating, dipping or spraying. Additional methods known in theart can also be utilized for this purpose. Methods of spraying includetraditional methods as well as microdeposition techniques with an inkjettype of dispenser. Additionally, a polymer can be deposited on animplantable device using photo-patterning to place the polymer on onlyspecific portions of the device. This coating of the device provides auniform layer around the device which allows for improved diffusion ofvarious analytes through the device coating.

In preferred embodiments of the invention, the inhibitory agent isformulated for release from the polymer coating into the environment inwhich the medical device is placed. Preferably, the inhibitory agent isreleased in a controlled manner over an extended time frame (e.g.,months) using at least one of several well-known techniques involvingpolymer carriers or layers to control elution. Some of these techniqueswere previously described in U.S. Patent Application 20040243225A 1.

Moreover, as described for example in U.S. Pat. No. 6,770,729, thereagents and reaction conditions of the polymer compositions can bemanipulated so that the release of the inhibitory agent from the polymercoating can be controlled. For example, the diffusion coefficient of theone or more polymer coatings can be modulated to control the release ofthe inhibitory agent from the polymer coating. In a variation on thistheme, the diffusion coefficient of the one or more polymer coatings canbe controlled to modulate the ability of an analyte that is present inthe environment in which the medical device is placed (e.g. an analytethat facilitates the breakdown or hydrolysis of some portion of thepolymer) to access one or more components within the polymer composition(and for example, thereby modulate the release of the inhibitory agentfrom the polymer coating). Yet another embodiment of the inventionincludes a device having a plurality of polymer coatings, each having aplurality of diffusion coefficients. In such embodiments of theinvention, the release of the inhibitory agent from the polymer coatingcan be modulated by the plurality of polymer coatings.

In yet another embodiment of the invention, the release of theinhibitory agent from the polymer coating is controlled by modulatingone or more of the properties of the polymer composition, such as thepresence of one or more endogenous or exogenous compounds, oralternatively, the pH of the polymer composition. For example, certainpolymer compositions can be designed to release a inhibitory agent inresponse to a decrease in the pH of the polymer composition.Alternatively, certain polymer compositions can be designed to releasethe inhibitory agent in response to the presence of hydrogen peroxide.

V. Methods of Treating Diseases Modulated by CCR6

In one aspect, the present invention provides methods of treating orpreventing a CCR6-mediated condition or disease by administering to asubject having such a condition or disease, a therapeutically effectiveamount of any compound of the invention. Preferred compounds for use inthe present methods are those compounds provided above as preferredembodiments, as well as compounds specifically exemplified in theExamples below, and provided with specific structures herein. The“subject” is defined herein to include animals such as mammals,including, but not limited to, primates (e.g., humans), cows, sheep,goats, horses, dogs, cats, rabbits, rats, mice and the like. Inpreferred embodiments, the subject is a human.

As used herein, the phrase “CCR6-mediated condition or disease” andrelated phrases and terms refer to a condition or disease characterizedby inappropriate, e.g., less than or greater than normal, CCR6functional activity. Inappropriate CCR6 functional activity might ariseas the result of CCR6 expression in cells which normally do not expressCCR6, increased CCR6 expression (leading to, e.g., inflammatory andimmunoregulatory disorders and diseases) or decreased CCR6 expression.Inappropriate CCR6 functional activity might also arise as the result ofCCL20 secretion by cells which normally do not secrete CCL20, increasedCCL20 expression (leading to, e.g., inflammatory and immunoregulatorydisorders and diseases) or decreased CCL20 expression. A CCR6-mediatedcondition or disease may be completely or partially mediated byinappropriate CCR6 functional activity. However, a CCR6-mediatedcondition or disease is one in which modulation of CCR6 results in someeffect on the underlying condition or disease (e.g., a CCR6 antagonistresults in some improvement in patient well-being in at least somepatients).

The term “therapeutically effective amount” means the amount of thesubject compound that will elicit the biological or medical response ofa tissue, system, animal or human that is being sought by theresearcher, veterinarian, medical doctor or other clinician.

Diseases and conditions associated with inflammation, infection andcancer can be treated or prevented with the present compounds andcompositions. In one group of embodiments, diseases or conditions,including chronic diseases, of humans or other species can be treatedwith inhibitors of CCR6 function. These diseases or conditions include:(1) allergic diseases such as systemic anaphylaxis or hypersensitivityresponses, drug allergies, insect sting allergies and food allergies,(2) inflammatory bowel diseases, such as Crohn's disease, ulcerativecolitis, ileitis and enteritis, (3) vaginitis, (4) psoriasis andinflammatory dermatoses such as dermatitis, eczema, atopic dermatitis,allergic contact dermatitis, urticaria and pruritus, Vitiligo (5)vasculitis, (6) spondyloarthropathies, (7) scleroderma, (8) asthma andrespiratory allergic diseases such as allergic asthma, allergicrhinitis, hypersensitivity lung diseases and the like, (9) autoimmunediseases, such as arthritis (including rheumatoid and psoriatic) as wellas for instance Hashimoto's thyroiditis and Grave's disease, multiplesclerosis, systemic lupus erythematosus, type I diabetes,glomerulonephritis, and the like, (10) graft rejection (includingallograft rejection and graft-v-host disease), and (11) other diseasesin which undesired inflammatory responses are to be inhibited, such asatherosclerosis, myositis, neurodegenerative diseases (e.g., Alzheimer'sdisease), encephalitis, meningitis, hepatitis, nephritis, sepsis,sarcoidosis, allergic conjunctivitis, otitis, chronic obstructivepulmonary disease, sinusitis, Behcet's syndrome and gout.

Preferably, the present methods are directed to the treatment ofdiseases or conditions selected from allergic diseases, psoriasis, skinconditions such as atopic dermatitis and asthma and scleroderma.

In another group of embodiments, modulation of CCR6 dependent regulatoryT cell trafficking may be modulated to treat diseases or conditionsincluding cancers, infectious diseases (viral infections, e.g., HIVinfection, and bacterial infections) and immunosuppressive diseases suchas organ transplant conditions and skin transplant conditions. The term“organ transplant conditions” is meant to include bone marrow transplantconditions and solid organ (e.g., kidney, liver, lung, heart, pancreasor combination thereof) transplant conditions.

Depending on the disease to be treated and the subject's condition, thecompounds of the present invention may be administered by oral,parenteral (e.g., intramuscular, intraperitoneal, intravenous, ICV,intracisternal injection or infusion, subcutaneous injection, orimplant), inhalation, nasal, vaginal, rectal, sublingual, or topicalroutes of administration and may be formulated, alone or together, insuitable dosage unit formulations containing conventional non-toxicpharmaceutically acceptable carriers, adjuvants and vehicles appropriatefor each route of administration. The present invention alsocontemplates administration of the compounds of the present invention ina depot formulation.

Those of skill in the art will understand that agents that modulate CCR6activity can be combined in treatment regimens with other therapeuticagents and/or with chemotherapeutic agents or radiation. In some cases,the amount of chemotherapeutic agent or radiation is an amount whichwould be sub-therapeutic if provided without combination with acomposition of the invention. Those of skill in the art will appreciatethat “combinations” can involve combinations in treatments (i.e., two ormore drugs can be administered as a mixture, or at least concurrently orat least introduced into a subject at different times but such that bothare in the bloodstream of a subject at the same time). Additionally,compositions of the current invention may be administered prior to orsubsequent to a second therapeutic regimen, for instance prior to orsubsequent to a dose of chemotherapy or irradiaition.

The compounds of the present invention are accordingly useful in theprevention and treatment of a wide variety of inflammatory andimmunoregulatory disorders and diseases.

In the treatment or prevention of conditions which require chemokinereceptor modulation an appropriate dosage level will generally be about0.001 to 100 mg per kg patient body weight per day which can beadministered in single or multiple doses. Preferably, the dosage levelwill be about 0.01 to about 25 mg/kg per day; more preferably about 0.05to about 10 mg/kg per day. A suitable dosage level may be about 0.01 to25 mg/kg per day, about 0.05 to 10 mg/kg per day, or about 0.1 to 5mg/kg per day. Within this range the dosage may be 0.005 to 0.05, 0.05to 0.5 or 0.5 to 5.0 mg/kg per day. For oral administration, thecompositions are preferably provided in the form of tablets containing1.0 to 1000 milligrams of the active ingredient, particularly 1.0, 5.0,10.0, 15.0. 20.0, 25.0, 50.0, 75.0, 100.0, 150.0, 200.0, 250.0, 300.0,400.0, 500.0, 600.0, 750.0, 800.0, 900.0, and 1000.0 milligrams of theactive ingredient for the symptomatic adjustment of the dosage to thepatient to be treated. The compounds may be administered on a regimen of1 to 4 times per day, preferably once or twice per day.

It will be understood, however, that the specific dose level andfrequency of dosage for any particular patient may be varied and willdepend upon a variety of factors including the activity of the specificcompound employed, the metabolic stability and length of action of thatcompound, the age, body weight, hereditary characteristics, generalhealth, sex and diet of the subject, as well as the mode and time ofadministration, rate of excretion, drug combination, and the severity ofthe particular condition for the subject undergoing therapy.

Diseases and conditions associated with inflammation, immune disorder,infection and cancer can be treated or prevented with the presentcompounds, compositions, and methods.

The compounds and compositions of the present invention can be combinedwith other compounds and compositions having related utilities toprevent and treat the condition or disease of interest, such asinflammatory or autoimmune disorders, conditions and diseases, includinginflammatory bowel disease, rheumatoid arthritis, osteoarthritis,psoriatic arthritis, polyarticular arthritis, multiple sclerosis,allergic diseases, psoriasis, atopic dermatitis and asthma, and thosepathologies noted above.

For example, in the treatment or prevention of inflammation orautimmunity or for example arthritis associated bone loss, the presentcompounds and compositions may be used in conjunction with ananti-inflammatory or analgesic agent such as an opiate agonist, alipoxygenase inhibitor, such as an inhibitor of 5-lipoxygenase, acyclooxygenase inhibitor, such as a cyclooxygenase-2 inhibitor, aninterleukin inhibitor, such as an interleukin-1 inhibitor, an NMDAantagonist, an inhibitor of nitric oxide or an inhibitor of thesynthesis of nitric oxide, a non steroidal anti-inflammatory agent, or acytokine-suppressing anti-inflammatory agent, for example with acompound such as acetaminophen, aspirin, codeine, fentanyl, ibuprofen,indomethacin, ketorolac, morphine, naproxen, phenacetin, piroxicam, asteroidal analgesic, sufentanyl, sunlindac, tenidap, and the like.Similarly, the instant compounds and compositions may be administeredwith an analgesic listed above; a potentiator such as caffeine, an H2antagonist (e.g., ranitidine), simethicone, aluminum or magnesiumhydroxide; a decongestant such as phenylephrine, phenylpropanolamine,pseudoephedrine, oxymetazoline, ephinephrine, naphazoline,xylometazoline, propylhexedrine, or levo desoxy ephedrine; anantitussive such as codeine, hydrocodone, caramiphen, carbetapentane, ordextromethorphan; a diuretic; and a sedating or non sedatingantihistamine.

Likewise, compounds and compositions of the present invention may beused in combination with other drugs that are used in the treatment,prevention, suppression or amelioration of the diseases or conditionsfor which compounds and compositions of the present invention areuseful. Such other drugs may be administered, by a route and in anamount commonly used therefor, contemporaneously or sequentially with acompound or composition of the present invention. When a compound orcomposition of the present invention is used contemporaneously with oneor more other drugs, a pharmaceutical composition containing such otherdrugs in addition to the compound or composition of the presentinvention is preferred. Accordingly, the pharmaceutical compositions ofthe present invention include those that also contain one or more otheractive ingredients or therapeutic agents, in addition to a compound orcomposition of the present invention. Examples of other therapeuticagents that may be combined with a compound or composition of thepresent invention, either administered separately or in the samepharmaceutical compositions, include, but are not limited to: (a) VLA-4antagonists, (b) corticosteroids, such as beclomethasone,methylprednisolone, betamethasone, prednisone, prenisolone,dexamethasone, fluticasone, hydrocortisone, budesonide, triamcinolone,salmeterol, salmeterol, salbutamol, formeterol; (c) immunosuppressantssuch as cyclosporine (cyclosporine A, Sandimmune®, Neoral®), tacrolirnus(FK-506, Prograf®), rapamycin (sirolimus, Rapamune®), Tofacitinib(Xeljanz®) and other FK-506 type immunosuppressants, and mycophenolate,e.g., mycophenolate mofetil (CellCept®); (d) antihistamines(H1-histamine antagonists) such as bromopheniramine, chlorpheniramine,dexchloipheniramine, triprolidine, clemastine, diphenhydramine,diphenylpyraline, tripelennamine, hydroxyzine, methdilazine,promethazine, trimeprazine, azatadine, cyproheptadine, antazoline,pheniramine pyrilamine, astemizole, terfenadine, loratadine, cetirizine,fexofenadine, descarboethoxyloratadine, and the like; (e) non steroidalanti asthmatics (e.g., terbutaline, metaproterenol, fenoterol,isoetharine, albuterol, bitolterol and pirbuterol), theophylline,cromolyn sodium, atropine, ipratropium bromide, leukotriene antagonists(e.g., zafmlukast, montelukast, pranlukast, iralukast, pobilukast andSKB-106,203), leukotriene biosynthesis inhibitors (zileuton, BAY-1005);(f) non steroidal anti-inflammatory agents (NSAIDs) such as propionicacid derivatives (e.g., alminoprofen, benoxaprofen, bucloxic acid,carprofen, fenbufen, fenoprofen, fluprofen, flurbiprofen, ibuprofen,indoprofen, ketoprofen, niroprofen, naproxen, oxaprozin, pirprofen,pranoprofen, suprofen, tiaprofenic acid and tioxaprofen), acetic acidderivatives (e.g., indomethacin, acemetacin, alclofenac, clidanac,diclofenac, fenclofenac, fenclozic acid, fentiazac, furofenac, ibufenac,isoxepac, oxpinac, sulindac, tiopinac, tolmetin, zidometacin andzomepirac), fenamic acid derivatives (e.g., flufenamic acid,meclofenamic acid, mefenamic acid, niflumic acid and tolfenamic acid),biphenylcarboxylic acid derivatives (e.g., diflunisal and flufenisal),oxicams (e.g., isoxicam, piroxicam, sudoxicam and tenoxican),salicylates (e.g., acetyl salicylic acid and sulfasalazine) and thepyrazolones (e.g., apazone, bezpiperylon, feprazone, mofebutazone,oxyphenbutazone and phenylbutazone); (g) cyclooxygenase-2 (COX-2)inhibitors such as celecoxib (Celebrex®) and rofecoxib (Vioxx®); (h)inhibitors of phosphodiesterase type IV (PDE IV); (i) gold compoundssuch as auranofin and aurothioglucose, (j) etanercept (Enbrel®), (k)antibody therapies such as orthoclone (OKT3), daclizumab (Zenapax®),basiliximab (Simulect®) and infliximab (Remicade®), adalimumab(Humira®), golimumab (Simponi®), rituximab (Rituxan®), tocilizumab(Actemra®), (1) other antagonists of the chemokine receptors, especiallyCCR5, CXCR2, CXCR3, CCR2, CCR3, CCR4, CCR7, CX₃CR1 and CXCR6; (m)lubricants or emollients such as petrolatum and lanolin, (n) keratolyticagents (e.g., tazarotene), (o) vitamin D₃ derivatives, e.g.,calcipotriene or calcipotriol (Dovonex®), (p) PUVA, (q) anthralin(Drithrocreme®), (r) etretinate (Tegison®) and isotretinoin and (s)multiple sclerosis therapeutic agents such as interferon β-1β(Betaseron®), interferon β-1α (Avonex®), azathioprine (Imurek®,Imuran®), glatiramer acetate (Capoxone®), a glucocorticoid (e.g.,prednisolone) and cyclophosphamide (t) DMARDS such as methotrexate andleflunomide (u) other compounds such as 5-aminosalicylic acid andprodrugs thereof, hydroxychloroquine; D-penicillamine; antimetabolitessuch as azathioprine, 6-mercaptopurine and methotrexate; DNA synthesisinhibitors such as hydroxyurea and microtubule disrupters such ascolchicine and proteasome inhibitors such as bortezomib (Velcade®). Theweight ratio of the compound of the present invention to the secondactive ingredient may be varied and will depend upon the effective doseof each ingredient. Generally, an effective dose of each will be used.Thus, for example, when a compound of the present invention is combinedwith an NSAID the weight ratio of the compound of the present inventionto the NSAID will generally range from about 1000:1 to about 1:1000,preferably about 200:1 to about 1:200. Combinations of a compound of thepresent invention and other active ingredients will generally also bewithin the aforementioned range, but in each case, an effective dose ofeach active ingredient should be used.

VI. Examples

The following examples are offered to illustrate, but not to limit theclaimed invention.

Reagents and solvents used below can be obtained from commercial sourcessuch as Aldrich Chemical Co. (Milwaukee, Wis., USA). ¹H-NMR wererecorded on a Varian Mercury 400 MHz NMR spectrometer. Significant peaksare provided relative to TMS and are tabulated in the order:multiplicity (s, singlet; d, doublet; t, triplet; q, quartet; m,multiplet) and number of protons. Mass spectrometry results are reportedas the ratio of mass over charge, followed by the relative abundance ofeach ion (in parenthesis). In tables, a single m/e value is reported forthe M+H (or, as noted, M−H) ion containing the most common atomicisotopes. Isotope patterns correspond to the expected formula in allcases. Electrospray ionization (ESI) mass spectrometry analysis wasconducted on a Hewlett-Packard MSD electrospray mass spectrometer usingthe HP1100 HPLC equipped with an Agilent Zorbax SB-C18, 2.1×50 mm, 5μcolumn for sample delivery. Normally the analyte was dissolved inmethanol at 0.1 mg/mL and 1 microliter was infused with the deliverysolvent into the mass spectrometer, which scanned from 100 to 1500daltons. All compounds could be analyzed in the positive ESI mode, usingacetonitrile/water with 1% formic acid as the delivery solvent. Thecompounds provided below could also be analyzed in the negative ESImode, using 2 mM NH₄OAc in acetonitrile/water as delivery system.

The following abbreviations are used in the Examples and throughout thedescription of the invention:

-   -   HPLC, High Pressure Liquid Chromatography; DMF, Dimethyl        formamide; TFA, Trifluoroacetic Acid; THF, Tetrahydrofuran;        EtOAc, Ethyl acetate; BOC₂O, di-tertbutyl dicarbonate or BOC        anhydride; HPLC, High Pressure Liquid Chromatography; DIPEA,        Diisopropyl ethylamine; HBTU,        O-(benzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium        hexafluorophosphate; dppf, 1,1′-Bis(diphenylphosphino)ferrocene;        Pd₂(dba)₃, Tris(dibenzylideneacetone)dipalladium(0); DIPEA,        diisopropylethylamine; DMP, dimethylphthalate; Me, methyl; Et,        ethyl; DCM, dichloromethane.

Compounds within the scope of this invention can be synthesized asdescribed below, using a variety of reactions known to the skilledartisan. One skilled in the art will also recognize that alternativemethods may be employed to synthesize the target compounds of thisinvention, and that the approaches described within the body of thisdocument are not exhaustive, but do provide broadly applicable andpractical routes to compounds of interest.

Certain molecules claimed in this patent can exist in differentenantiomeric and diastereomeric forms and all such variants of thesecompounds are claimed.

The detailed description of the experimental procedures used tosynthesize key compounds in this text lead to molecules that aredescribed by the physical data identifying them as well as by thestructural depictions associated with them.

Those skilled in the art will also recognize that during standard workup procedures in organic chemistry, acids and bases are frequently used.Salts of the parent compounds are sometimes produced, if they possessthe necessary intrinsic acidity or basicity, during the experimentalprocedures described within this patent.

Example 1: Synthesis of Methyl5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)quinoline-8-carboxylate

(a) To a stirred solution of H₂O (80 mL) and H₂SO₄ (120 mL) at 0° C. wasadded 5-bromo-8-methylquinoline (25 g, 112.6 mmol). After obtaining asolution, CrO₃ was introduced (16 g, 157.6 mmol) in portion wise whilemaintaining the internal temperature at 70° C. The reaction mixture wasstirred for 1 h at 70° C. An additional CrO₃ (16 g, 157.6 mmol) wasadded in portions and stirred at 80° C. for 2.5 h. After completion ofthe reaction, it was cooled to r.t, poured onto crushed ice, neutralizedwith aqueous ammonium hydroxide to get the solids. The solids werefiltered, dried under high vacuum for 16 h to get the crude5-bromoquinoline-8-carboxylic acid (28.0 g) as a green colored solid.

(b) K₂CO₃ (61.3 g, 444.0 mmol) and methyl iodide (63.1 g, 444.0 mmol)were added to a stirred suspension of 5-bromoquinoline-8-carboxylic acid(28.0 g, 111.0 mmol) in DMF (250 mL) at r.t. The reaction mixture washeated at 45° C. for 36 h, cooled to r.t, filtered the solids, washedwith ethyl acetate (100 mL). The filtrate was diluted with water,extracted with ethyl acetate (3×300 mL) and washed with water (3×100mL). The ethyl acetate layer was dried (Na₂SO₄), filtered andconcentrated under reduced pressure. The residue was purified by flashcolumn chromatography on silica gel, eluting with ethyl acetate inhexanes (0-20%) to afford methyl-5-bromoquinoline-8-carboxylate as acream color solid (20.5 g, 70% for 2 steps). ¹H NMR (400 MHz, CDCl₃) δ9.07 (dd, J=4.3, 1.5 Hz, 1H), 8.60 (dd, J=8.7, 1.6 Hz, 1H), 7.88 (s,2H), 7.58 (dd, J=8.6, 4.0 Hz, 1H), 4.05 (s, 3H)

(c) A solution of methyl-5-bromoquinoline-8-carboxylate (20.5 g, 77.06mmol),4,4,5,5-tetramethyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3,2-dioxaborolane(21.4 g, 84.7 mmol), and KOAc (18.8 g, 192.6 mmol) in dry 1,4-dioxane(200 mL) was purged with nitrogen gas for 10 min. Then, Pd(dppf)Cl₂(3.14 g, 7.70 mmol) was added and the resulting mixture was heated at95° C. for 8 h, cooled to r.t and the excess solvent was removed undervacuum. The residue was diluted with ether, filtered through a Celiteplug and washed with ether (200 mL); the filtrate was concentrated underreduced pressure. The resulting residue was purified by columnchromatography on silica gel, eluting with 0-50% ethyl acetate inhexanes to afford a red color thick syrup/solid which was dissolved inether and cooled to −20° C. and stored for 12 h, the separated lightpink color solid was filtered and dried to get pure methyl5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl) quinoline-8-carboxylate(17.5 g, 78%): ¹H NMR (400 MHz, CDCl₃) δ 9.14 (dd, J=8.6, 0.6 Hz, 1H),9.02-9.01 (m, 1H), 8.14 (d, J=7.0 Hz, 1H), 7.94 (d, J=7.0 Hz, 1H), 7.48(dd, J=8.6, 4.3 Hz, 1H), 4.06 (s, 3H), 1.43 (s, 12H).

Example 2: Synthesis of 3,4-dichloro-2-iodoaniline

To a stirred suspension of 3,4-dichloroaniline (20 g, 123 mmol), HI(48%, 15.7 g, 123 mmol), H₂O₂ (30%, 8.3 g, 246 mmol) in H₂O (62 mL) atr.t. The reaction mixture was stirred in dark at r.t for overnight. Thesupernatant was discarded, diluted with ethyl acetate:hexanes (1:10),quenched with saturates NaHSO₃, stirred for 1 h at ambient temperature,filtered the solids. The filtrate was washed with water, saturatedaqueous NaHCO3, dried (Na2SO4), filtered and concentrated to get 1:4regioisomeric mixture of iodo derivative (minor isomer is required). Itwas recrystallized from cyclohexane to afford 1:1 mixture ofregioisomers enriched in the filtrate. This mixture was further purifiedby flash column chromatography on silica gel, eluting with ethylacetate:hexanes (0-2%) to get the required compound as a cream colorsolid (5.2 g, ˜15% yield). ¹H NMR (400 MHz, CDCl₃) δ 7.22 (d, J=8.6 Hz,1H), 6.60 (d, J=8.6 Hz, 1H), 4.32 (br, 2H).

Example 3: 3-chloro-2-iodo-4-(trifluoromethoxy)aniline

The title compound was synthesized in a similar fashion to Example 2 in18% yield. ¹H NMR (400 MHz, CDCl₃) δ 7.11 (d, J=8.9 Hz, 1H), 6.67 (d,J=8.9 Hz, 1H), 4.30 (br, 2H).

Example 4: Synthesis of 2-bromo-3-fluoro-4-trifluoromethoxyaniline

(a) In a 20 mL vial was added 3-fluoro-4-trifluoromethoxyaniline (150mg, 0.769 mmol) in acetic acid (0.5 mL) at room temperature. Bromine(118 μL, 2.30 mmol, 3 eq.) was added and the reaction was stirred forseveral hours. Dichloromethane (2 mL) was added to break up thesolidified reaction mixture, followed by more bromine (40 μL, 0.781mmol, 1 eq.) addition. After stirring for another hour, dichloromethanewas removed by gently blowing a stream of nitrogen to the reactionmixture. The solid was filtered and washed thoroughly with water andthen dried under vacuum to give 2,6-dibromo-3-fluoro-4-trifluoromethoxyaniline as a white solid (172 mg,0.489 mmol, 64% yield).

(b) In a 40 mL vial was added 2,6-dibromo-3-fluoro-4-trifluoromethoxyaniline (172 mg, 0.489 mmol),followed by tin(II) chloride dihydrate (122 mg, 0.540 mmol, 1.1 eq.).Acetic acid (725 μL) and concentrated hydrochloric acid (580 μL) wasadded to make a solution. The reaction mixture was heated at 115° C. for1.5 hours. After cooling the reaction to room temperature, most ofacetic acid was removed under vacuum. Water was added and the productwas extracted with dichloromethane three times. The organic layer wasdried over anhydrous sodium sulfate. After removing the solvent underreduced pressure, the crude material was purified using silica gelcolumn chromatography. The recovered 2,6-dibromo-3-fluoro-4-trifluoromethoxyaniline was eluted using a gradientof 5-10% ethyl acetate in hexanes followed by the desired product,2-bromo-3-fluoro-4-trifluoromethoxyaniline, which was eluted with 15%ethyl acetate in hexanes (64.8 mg, 0.236 mmol, 48%). ¹H NMR (400 MHz,CDCl₃) δ 7.06 (dd, J=9.0, 9.0 Hz, 1H), 6.52 (dd, J=9.0, 2.0 Hz, 1H),4.27 (br, 2H).

Example 5: Synthesis of4-chloro-5-bromo-6-amino-2,2-difluoro-1,3-benzodioxole

(a) To a stirred solution of 4-amino-2,2-diflouro-1,3-benzodioxole (2.5g, 14.4 mmol) in dichloromethane at 0° C. was added N-bromosuccinimide(2.7 g, 15.2 mmol) portion wise. The solution was stirred at 0° C. for30 min, then at room temperature for 1 h. The reaction mixture wasquenched with IM sodium thiosulfate solution and diluted with deionizedwater, extracted with dichloromethane (2×50 mL). The combined organiclayers were dried (Na₂SO₄), filtered, and concentrated in vacuo. Thecrude product was purified by flash chromatography (SiO₂, 0-20% ethylacetate in hexanes) to recover starting material 1 g and afford5-bromo-4-amino-2,2-diflouro-1,3-benzodioxole as colorless oil (2.33 g,9.25 mmol, 64%). MS: (ES) m/z calculated for C₇H₄BrF₂NO [M+H]⁺ 252.0,found 252.

(b) To a stirred solution of copper (II) chloride (2.49 g, 18.5 mmol)and tert-butyl nitrite (2.39 g, 23.13 mmol) in acetonitrile at 55° C.was added a solution of 5-bromo-4-amino-2,2-diflouro-1,3-benzodioxole(2.33 g, 9.25 mmol) in acetonitrile. The solution was stirred at 55° C.for 30 min, then cool down to room temperature. The reaction mixture wasquenched with 5% hydrochloric acid solution and diluted with deionizedwater, extracted with ethyl acetate (2×50 mL). The organic layers weredried (Na₂SO₄), filtered, and concentrated in vacuo. The crude productwas purified by flash chromatography (SiO₂, 0-20% ethyl acetate inhexanes) to afford 5-bromo-4-chloro-2,2-diflouro-1,3-benzodioxole asyellowish oil (1.5 g, 5.53 mmol, 60%). MS: (ES) m/z calculated forC₇H₂BrClF₂O₂[M+H]⁺ 272.0, no MS found.

(c) The solution of 5-bromo-4-chloro-2,2-diflouro-1,3-benzodioxole (1.5g, 5.53 mmol) in 10 mL conc. sulfuric acid was cooled to −10° C. Then asolution of 1 mL fuming nitric acid and 2 mL of conc. sulfuric acid wasadded drop wise. The resulting solution was stirred at −10° C. for ˜2 h.Then the reaction mixture was poured into ice water and a yellow solidprecipitated out. Filtered off the solid, rinsed with water andcollected the solid. The crude solid was purified by flashchromatography (SiO₂, 0-5% ethyl acetate in hexanes) to afford pure6-nitro-5-bromo-4-chloro-2,2-diflouro-1,3-benzodioxole (1.0 g, 3.16mmol, 57%). MS: (ES) m/z calculated for C₇HBrClF₂NO₄ [M+H]⁺ 316.0, found316.

(d) To the slurry of6-nitro-5-bromo-4-chloro-2,2-diflouro-1,3-benzodioxole (1.0 g, 3.16mmol) in ethanol (4 mL) and water (1 mL) was added ammonium chloride(3.38 g, 63.2 mmol) and iron powder (1.06 g, 18.96 mmol). The mixturewas warmed up to 90° C. for 1 h and then cooled down. Filtered through aCelite plug and concentrated in vacuo. The crude product was purified byflash chromatography (SiO₂, 0-30% ethyl acetate in hexanes) to afford6-amino-5-bromo-4-chloro-2,2-diflouro-1,3-benzodioxole as a white solid(0.90 g, 3.16 mmol, 100%). MS: (ES) m: calculated for C₇H₃BrClF₂NO₂[M+H]⁺ 286.0, found 286.

Example 6: Synthesis of2,2-difluoro-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzo-[d][1,3l]dioxol-5-amine

The title compound was synthesized in a similar fashion to Example 1 in550% yield. ¹H NMR (400 MHz, CDCl₃) δ 7.22 (s, 1H), 6.31 (s, 1H), 4.80(br, 2H), 1.34 (s, 12H).

Example 7: Synthesis of2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-4-(trifluoromethoxy)aniline

The title compound was synthesized in a similar fashion to Example 1. ¹HNMR (400 MHz, CDCl₃) δ 7.43 (br, 1H), 7.10 (dd, J=4.2, 1.8 Hz, 1H), 6.55(d, J=7.1 Hz, 1H), 1.34 (s, 12H).

Example 8: Synthesis of5-[6-[(4-tert-butyl-3-fluorophenyl)sulfonylamino]-2,2-difluoro-1,3-benzodioxol-5-yl]quinoline-8-carboxylicacid

(a) To a solution of2,2-difluoro-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3-benzodioxol-5-amine(Example 5, 2.7 g, 9.03 mmol) in dry pyridine (12 mL) was added the4-tert-butyl-3-fluorophenylsulfonyl chloride (2.82 g, 11.28 mmol) andthe reaction mixture was heated at 80° C. for overnight. It was cooledto r.t. excess solvent was removed in vacuo. The residue was dilutedwith saturated aqueous ammonium chloride, extracted with DCM, purifiedby flash column using hexanes: ethyl acetate mixture as an eluent onsilica column to get the pure compound (3.28 g, 71%)

(b) A mixture of the4-tert-butyl-N-[2,2-difluoro-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3-benzodioxol-5-yl]-3-fluorobenzenesulfonamide(3.2 g, 6.22 mmol), methyl-5-bromoquinoline-8-carboxylate (Example 1,step-b, 1.66 g, 6.22 mmol) and 2 M aq. K₂CO₃ (7.8 mL, 15.55 mmol) in1,4-dioxane (50 mL) was purged with nitrogen for 5 minutes. Pd(PPh₃)₄(360 mg, 0.31 mmol, 5 mol %) was added and the reaction mixture washeated at 95° C. for overnight. The reaction mixture was cooled to r.t.LiOH.H₂O (2.6 g, 62.2 mmol) was added and heated at 80° C. for 1 hour.Cooled to r.t, pH of the reaction medium was adjusted to ˜6 with 2Naq.HCl, extracted with ethyl acetate, dried (Na₂SO₄), filtered andconcentrated. The residue was triturated with methanol to get pure(>95%) title compound as an off-white solid (1.75 g, 50%). ¹H NMR (400MHz, CD₃OD) δ 8.96-8.94 (m, 1H), 8.57 (d, J=7.4 Hz, 1H), 7.93-7.91 (m,1H), 7.57-7.54 (m, 2H), 7.42- (t, J=7.8 Hz, 1H), 7.30-7.26 (m, 1H),7.18-7.15 (m, 1H), 7.05 (d, J=7.5 Hz, 1H), 6.88 (s, 1H), 6.56 (br, 1H),1.43 (s, 9H); MS: (ES) m/z calculated for C₂₇H₂₁F₃N₂O₆S.

Example 9: Synthesis of5-[2-[(4-tert-butyl-3-fluorophenyl)sulfonylamino]-5-(trifluoromethoxy)phenyl]quinoline-8-carboxylicacid

The title compound was synthesized in a similar fashion to Example 8. ¹HNMR (400 MHz, CD₃OD) δ 9.02 (dd, J=4.4, 1.6 Hz, 1H), 8.60 (d, J=7.5 Hz,1H), 8.07 (dd, J=8.6, 1.7 Hz, 1H), 7.67 (dd, J=8.7, 4.4 Hz, 1H), 7.56(d, J=8.9 Hz, 1H), 7.47 (ddt, J=9.0, 1.9, 1.1 Hz, 1H), 7.39-7.24 (m,3H), 7.14 (dd, J=8.3, 2.0 Hz, 1H), 7.03 (dd, J=12.0, 2.0 Hz, 1H), 1.39(s, 9H); MS: (ES) m/z calculated for C₂₇H₂₂F₄N₂O₅S [M+H]⁺ 558.12 found558.1.

Example 10:5-[2-[(4-tert-butyl-3-fluorophenyl)sulfonylamino]-5-chlorophenyl]quinoline-8-carboxylicacid

The title compound was synthesized in a similar fashion to Example 8. ¹HNMR (400 MHz, CD₃OD) δ 9.10 (dd, J=4.7, 1.6 Hz, 1H), 8.66 (d, J=7.8 Hz,1H), 8.27 (dd, J=8.6, 1.6 Hz, 1H), 7.78 (dd, J=8.6, 4.7 Hz, 1H), 7.54(dd, J=8.6, 2.7 Hz, 1H), 7.41-7.36 (m, 3H), 7.16 (dd, J=8.2, 1.9 Hz,1H), 7.06 (dd, J=11.1, 1.9 Hz, 1H), 1.40 (s, 9H); MS: (ES) m/zcalculated for C₂₆H₂₂ClFN₂O₄S [M+H]⁺ 513.1 found 513.1.

Example 11: Synthesis of5-[6-[(4-tert-butyl-3-fluorophenyl)sulfonylamino]-2,3-dichlorophenyl]quinoline-8-carboxylicacid

(a) To a solution of 3,4-dichloro-2-iodoaniline (example 2, 2.2 g, 7.67mmol) in dry pyridine (8 mL) was added4-tert-butyl-3-fluorophenylsulfonyl chloride (2.11 g, 8.43 mmol) and thereaction mixture was heated at 80° C. for overnight. LCMS indicatedpresence of mono and bis-sulfonamides. 10 N aq. NaOH (3 mL) and ethanol(2 mL) were added to hydrolyze the bis-sulfonamide. The reaction mixturewas then heated at 80° C. for 2 h. It was cooled to r.t, excess solventwas removed in vacuo to afford a dark brown solid, diluted with DCM,washed with 1N aq.HCl, and purified by flash column using hexanes: ethylacetate mixture as an eluent on silica column to get the pure4-tert-butyl-N-(3,4-dichloro-2-iodophenyl)-3-fluorobenzsulfonamide as anoff white solid (3.5 g, 91%)

(b) A mixture of4-tert-butyl-N-(3,4-dichloro-2-iodophenyl)-3-fluorobenzsulfonamide (9 g,17.9 mmol), Methyl5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)quinoline-8-carboxylate(7.8 g, 25.1 mmol), K₃PO₄.H₂O (10.3 g, 44.75 mmol) and SPhos (0.73 g,1.79 mmol) in n-BuOH:H₂O (75:25 mL) solvent system was purged withnitrogen for 15 minutes. Then Pd₂(dba)₃ (0.65 g, 0.72 mmol) was addedand heated at 85° C. for 6 h, cooled to r.t, filtered through a Celiteplug, washed with ethyl acetate and the filtrate was concentrated invacuo. The crude product was diluted with THF/H₂O (60/10 mL) and lithiumhydroxide monohydrate (7.52 g, 179 mmol) was added. The reaction mixturewas then heated at 60° C. for overnight, cooled to r.t, the pH wasadjusted to ˜6 by 2N aq. HCl, extracted with ethyl acetate (3×500 mL),the combined ethyl acetate layer was washed with water, brine, dried(Na₂SO₄), filtered and concentrated. The residue was purified by flashcolumn using ethyl acetate and hexanes as eluents to get the productwhich was further triturated from methanol to afford the title compoundas an off-white solid (7.2 g, 73%). ¹H NMR (400 MHz, CDCl₃) δ 8.98 (dd,J=4.4, 1.6 Hz, 1H), 8.69 (d, J=7.5 Hz, 1H), 7.77-7.69 (m, 2H), 7.63 (d,J=9.0 Hz, 1H), 7.52 (dd, J=8.6, 4.3 Hz, 1H), 7.42 (t, J=8.0 Hz, 1H),7.32 (dd, J=4.4, 1.6 Hz, 1H), 7.17 (dd, J=11.4, 2.0 Hz, 1H), 7.05 (d,J=7.5 Hz, 1H), 6.30 (s, 1H), 1.48 (s, 9H); MS: (ES) m/z calculated forC₂₆H₂₁FN₂O₄S [M+H]; 547.06 found 547.1.

Example 12:5-[2-[(4-tert-butyl-3-fluorophenyl)sulfonylamino]-5-cyanophenyl]quinoline-8-carboxylicacid bis sodium salt

The title compound was synthesized in a similar fashion to Example 11.

0.1 N aq. NaOH (2 eq) was added to the free acid suspended inacetonitrile and water and lyophilized to get the bis sodium salt. ¹HNMR (400 MHz, CD₃OD) δ 9.10 (dd, J=4.8, 1.6 Hz, 1H), 8.69 (d, J=7.5 Hz,1H), 8.19 (dd, J=8.7, 1.7 Hz, 1H), 7.89 (dd, J=8.5, 2.1 Hz, 1H),7.81-7.68 (m, 3H), 7.51-7.38 (m, 2H), 7.32 (dd, J=8.3, 2.0 Hz, 1H), 7.22(dd, J=11.9, 2.0 Hz, 1H), 1.41 (s, 9H); MS: (ES) m/z calculated forC₂₇H₂₂FN₃O₄S [M+H]⁺ 504.13 found 504.

Example 13: Synthesis of5-[6-[(4-tert-butyl-3-fluorophenyl)sulfonylamino]-2-fluoro-3-(trifluoromethoxy)phenyl]quinoline-8-carboxylicacid

The title compound was synthesized in a similar fashion to Example 11.¹H NMR (400 MHz, CDCl₃) δ 9.00 (dd, J=4.3, 1.7 Hz, 1H), 8.72 (d, J=7.5Hz, 1H), 7.86 (dt, J=8.6, 1.5 Hz, 1H), 7.64-7.54 (m, 2H), 7.53-7.33 (m,3H), 7.30-7.18 (m, 2H), 1.42 (s, 9H); MS: (ES) m/z calculated forC₂₇H21N₂O₅SF₅ [M−H]⁻ 579.1, found 579.1.

Example 14: Synthesis of5-[6-[(4-tert-butylphenyl)sulfonylamino]-2-chloro-3-(trifluoromethoxy)phenyl]quinoline-8-carboxylicacid bis sodium salt

The title compound was synthesized in a similar fashion to Example 11.¹H NMR (400 MHz, CD₃OD) δ 8.79-8.77 (m, 1H), 7.72 (d, J=7.4 Hz, 1H),7.57-7.54 (m, 1H), 7.52 ((d, J=9.4 Hz, 1H), 7.38-7.35 (m, 2H), 7.32-7.29(m, 2H), 7.23-7.18 (m, 2H), 6.99 (d, J=7.4 Hz, 1H), 1.32 (s, 9H); MS:(ES) m/z calculated for C₂₇H₂₂ClF₃N₂O₅S [M+H]⁻ 579.09, found 579.1.

Example 15: Synthesis of5-[6-[(4-tert-butylphenyl)sulfonylamino]-2-chloro-3-(trifluoromethoxy)phenyl]quinoline-8-carboxylicacid

The title compound was synthesized in a similar fashion to Example 11.¹H NMR (400 MHz, CD₃OD 9.06 (dd, J=4.8, 1.6 Hz, 1H), 8.61 (d, J=8.0 Hz,1H), 7.94 (dd, J=8.8, 1.6 Hz, 1H), 7.72-7.60 (m, 3H), 7.42 (t, J=8.1 Hz,1H), 7.20 (dd, J=8.0, 1.6 Hz, 1H), 7.16-7.06 (m, 2H), 1.42 (s, 9H); MS:(ES) m/z calculated for C₂₇H₂₁ClF₄N₂O₅S [M+H]⁺ 597.08 found 597.1.

Example 16: Synthesis of5-[6-[(4-tert-butyl-3-fluorophenyl)sulfonylamino]-4-chloro-2,2-difluoro-1,3-benzodioxol-5-yl]quinoline-8-carboxylicacid

The title compound was synthesized in a similar fashion to Example 11.¹H NMR (400 MHz, CD₃OD) δ 9.08 (dd, J=4.7, 1.6 Hz, 1H), 8.58 (d, J=7.4Hz, 1H), 8.11 (dd, J=8.6, 1.6 Hz, 1H), 7.72 (dd, J=8.6, 4.7 Hz, 1H),7.43-7.39 (m, 2H), 7.18 (dd, J=8.2, 2.0 Hz, 1H), 7.08 (dd, J=10.2, 2.0Hz, 1H), 1.40 (s, 9H); MS: (ES) m/z calculated for C₂₇H₂₀ClF₃N₂O₆S[M+H]⁻ 593.07 found 593.1.

Example 17: Synthesis of4-tert-butyl-N-[2,2-difluoro-6-[8-(1H-tetrazol-5-yl)-5-quinolyl]-1,3-benzodioxol-5-yl]-3-fluorobenzenesulfonamide

(a) A mixture of 5-bromo-8-iodoquinoline (0.52 g, 1.55 mmol), Zn(CN)₂(218 mg, 1.86 mmol), and dppf (103 mg, 0.186 mmol) in DMF (2.5 mL) waspurged with N₂ (gas) for 5 minutes. Pd₂(dba)₃ (85 mg, 0.093 mmol) wasadded and the resulting mixture was heated at 95° C. for 5 h. Aftercompletion of the reaction, it was cooled to r.t., diluted with water,and the aqueous solution was extracted with ethyl acetate (3×50 mL). Thecombined organic layer was washed with brine, dried over Na₂SO₄ andconcentrated under reduced pressure. The crude product was purified bycolumn chromatography on silica gel, eluting with ethyl acetate-hexanes(5-25%) to get 5-bromoquinoline-8-carbonitrile as a brown solid (72 mg).

(b) The Suziki reaction was carried out similar to Example 8. 72 mg ofthe starting material afforded 70 mg of the product.

(c) To a stirred solution of the above nitrile (35 mg, 0.064 mmol) inH₂O/IPA (5:1, 2 mL) at r.t were added ZnBr₂ (43 mg, 0.324 mmol) and NaN₃(21 mg, 0.324 mmol). The reaction mixture was heated at 100° C. for 16h. After completion of the reaction, it was cooled to r.t andneutralized with 1N aq. HCl. The aqueous solution was extracted withethyl acetate (2×25 mL) and the combined organic layers were washed withbrine, dried over Na₂SO₄, filtered and concentrated under reducedpressure. The crude product was triturated with methanol to give thetitle compound as a brownish solid (8 mg). ¹H NMR (400 MHz, DMSO-d) δ9.84 (s, 1H), 9.04 (dd, J=4.2, 1.7 Hz, 1H), 8.45 (d, J=7.5, Hz, 1H),7.87 (dd, J=8.6, 1.7 Hz, 1H), 7.58 (dd, J=8.6, 4.2 Hz, 1H), 7.43 (d,J=12.6 Hz, 1H), 7.24 (t, J=8.1, Hz, 1H), 7.16-7.0 (m, 3H), 1.24 (s, 9H);MS: (ES) m/z calculated for C₂₈H₂₂F₃N₅O₄S [M+H]⁺ 582.13 found 582.1.

Example 18: Synthesis of5-[2-[(4-tert-butylphenyl)sulfonylamino]-5-cyanophenyl]quinoline-8-carboxylicacid

The title compound was synthesized in a similar fashion to Example 11.¹H NMR (400 MHz, CDCl₃) δ 9.00 (dd, J=4.3, 1.6 Hz, 1H), 8.68 (d, J=7.4Hz, 1H), 7.92 (d, J=8.6 Hz, 1H), 7.87 (dd, J=8.6, 1.2 Hz, 1H), 7.78 (dd,J=9.0, 2.0 Hz, 1H), 7.62-7.50 (m, 5H), 7.45 (d, J=7.9 Hz, 1H), 7.19 (d,J=7.5 Hz, 2H), 6.67 (s, 1H), 1.37 (s, 9H); MS: (ES) m/z calculated forC₂₇H23N₃O₄S [M+H]⁺ 486.14 found 486.1.

Example 19: Synthesis of5-[6-[(4-tert-butylphenyl)sulfonylamino]-2-chloro-3-cyanophenyl]quinoline-8-carboxylicacid trifluoroacetic acid salt

The title compound was synthesized in a similar fashion to Example 11.¹H NMR (400 MHz, DMSO-d6) δ 9.85 (bs, 1H), 9.12-9.13 (m, 1H), 8.48 (d,J=7.4 Hz, 1H), δ 8.06 (bs, 1H), 7.89-7.87 (m, 1H), 7.70-7.52 (m, 7H),7.16 (d, J=6.7 Hz, 1H), 1.30 (s, 9H); MS: (ES) m/z calculated forC₂₇H₂₂ClN₃O₄S [M+H]⁻ 520.09, found 520.1.

Example 20: Synthesis of5-[6-[(4-tert-butylphenyl)sulfonylamino]-3-cyano-2-fluorophenyl]quinoline-8-carboxylicacid hydrochloric acid salt

The title compound was synthesized in a similar fashion to Example 11.¹H NMR (400 MHz, CDCl₃ 9.04 (dd, J=4.3, 1.6 Hz, 1H), 8.76 (d, J=7.4 Hz,1H), 7.84-7.75 (m, 3H), 7.64-7.53 (m, 5H), 7.22 (d, J=7.4 Hz, 1H), 6.59(bs, 1H) 1.37 (s, 9H); MS: (ES) m/z calculated for C₂₇H₂₂FN₃O₄S [M+H]⁺504.1 found 504.4.

Biological Example 1: Ligand Binding Assay

Ligand binding assay was used to determine the ability of potential CCR6antagonists to block the interaction between CCR6 and its ligand CCL20(MIP3alpha). L1.2 cells stably expressing the CCR6 receptor, werecentrifuged and resuspended in assay buffer (20 mM HEPES pH 7.1, 140 mMNaCl, 1 mM CaCl₂, 5 mM MgCl₂, 0.1% sodium azide and with 0.1% bovineserum albumin) to a concentration of 5×10^5 cells/mL. Binding assayswere set up as follows. First, 0.1 mL of cells (5×104 cells/well) wasadded to the assay plates containing the compounds, giving a finalconcentration of ˜2-10 uM each compound for screening (or part of a doseresponse for compound IC50 determinations). Then 0.1 mL of ¹²⁵I labeledCCL20 (obtained from PerkinElmer; Waltham, Mass.) diluted in assaybuffer to a final concentration of ˜50 pM, yielding ˜30,000 cpm perwell, was added, the plates sealed and incubated for approximately 3hours at 25° C. on a shaker platform. Reactions were aspirated onto GF/Bglass filters pre-soaked in 0.3% polyethyleneimine (PEI) solution, on avacuum cell harvester (Packard Instruments; Meriden, Conn.).Scintillation fluid (50 uL; Microscint 20, Packard Instruments) wasadded to each well, the plates were sealed and radioactivity measured ina Top Count scintillation counter (Packard Instruments). Control wellscontaining either diluent only (for total counts) or 20 uM compound wereused to calculate the percent of total inhibition for compound. Thecomputer program Prism from GraphPad, Inc. (San Diego, Ca) was used tocalculate IC50 values. IC50 values are those concentrations required toreduce the binding of labeled TARC to the receptor by 50%. Compounds inFIG. 1 having an IC₅₀ value in the binding assay of less than 100 nM arelabeled (+++); from 100-500 nM are labeled (++); and less than or equalto 20 μM but above 500 nM are labeled (+).

Biological Example 2: Migration/Chemotaxis Assay

A serum chemotaxis assay was used to determine the efficacy of potentialreceptor antagonists at blocking the migration mediated throughchemokine receptors, such as CCR6. This assay was routinely performedusing the ChemoTX® microchamber system with a 5-μm pore-sizedpolycarbonate membrane. To begin such an assay, chemokine-receptorexpressing cells (such as L1.2 cells stably expressing CCR6) werecollected by centrifugation at 400×g at room temperature, then suspendedat 50 million/ml in human serum. The compound being tested or anequivalent volume of its solvent (DMSO) was then added to the cell/serummixture at a final DMSO concentration of 0.25% (v/v). Separately,recombinant human CCL20 was diluted with chemotaxis buffer (HBSS+0.1%BSA), generally spanning a range from 0.01 nM to 500 nM, after which 29μl of diluted chemokine was placed in the lower wells of the ChemoTX®plate. The 5-μm (pore size) polycarbonate membrane was placed onto theplate, and 20 μL of the cell/compound mixture was transferred onto eachwell of the membrane. The plates were incubated at 37° C. for 90minutes, after which the polycarbonate membranes were removed and 5 μlof the DNA-intercalating agent CyQUANT (Invitrogen, Carlsbad, Calif.)was added to the lower wells. The amount of fluorescence, correspondingto the number of migrated cells, was measured using a Spectrafluor Plusplate reader (TECAN, San Jose, Calif.).

(a) Evaluation of a Compound of Interest in an Oxazalone Induced Modelof DTH

Compounds of the invention were assessed in the murine model of dermaldelayed type hypersensitivity (DTH) induced by oxazolone. Briefly, 8-10week old BALB/c mice were sensitized topically with a 1% solution ofoxazolone dissolved in ethanol on their shaved abdomens on day 0. On day6 post sensitization mice were dosed orally with either vehicle orincreasing doses of compounds 1.061 and 1.033 of the inventionimmediately prior to and 4 hours following a topical challenge with a0.5% solution of oxazolone in ethanol on the right ear. The followingday (day 7), ear thicknesses were measured using caliper measurements.Animals treated with compound had significantly reduced ear swellingcompared to vehicle treated controls indicating a compound mediateddecrease in oxazolone induced dermal hypersensitivity.

(b) Evaluation of a Compound of Interest in an Imiquimod Induced Modelof Psoriasis

Multiple mouse models of psoriasis are known. For example, in theImiquimod-induced mouse model of psoriasis, hair was removed from theback of balb/c mice three days prior to the application of 5% Imiquimodonce a day. Vehicle (1% HPMC) or CCR6 compound was administered eg.orally at between 5 and 200 mg/kg once/twice daily at the initiation ofImiquimod application and continued for five to ten days. Therapeuticvalue was assessed using the following methodologies: daily electroniccaliper measurements and immunohistochemistry at the end point to assessskin thickening, flow cytometry and immunofluorescence at the end pointto measure leukocyte infiltration and inflammation, and mRNAquantitation via Luminex at days two and five to assess molecularchanges. Treatment of animals with compound 1.033 or 1.061 of theinvention resulted in a significant improvement in disease.

(c) Evaluation of a Compound of Interest in an IL-23 Induced Model ofPsoriasis.

Another method to induce phenotypic changes associated with psoriasisinvolved intra-dermal injections of IL-23. Briefly, 500 ng ofrecombinant murine IL-23 was intra-dermally injected every other dayinto the right ear of C57BL6/N mice for a total of 5-6 administrationsover a period of ten to twelve days. Concomitantly, PBS wasintra-dermally injected every other day into the left ear of the samemice. CCR6 compound was dosed prophylactically at eg. between 5-200mg/kg once/twice daily via the oral or other route. Therapeutic dosingwas performed by orally providing IL-23 challenged mice with vehiclecontrol for four days. On the fifth day, animals were dosed with acompound of the invention at eg. 5-200 mg/kg once/twice daily via theoral or other route (at this point, two rounds of IL-23 have beenintra-dermally injected into the right ear). Efficacy was assessedquantitatively by manually measuring ear thickness using a caliperdaily. Treatment of animals with compound 1.033 or 1.061 of theinvention resulted in a significant improvement in disease.

(d) Evaluation of a Compound of Interest in a Mouse Model of SepticShock.

This example describes a procedure to evaluate the efficacy of CCR6antagonists for treatment of septic shock. An animal model of endotoxicshock can be induced by injecting rodents with lipopolysaccharide (LPS).Three series of mouse groups, comprising 15 mice per group, are treatedwith an intra-peritoneal injection of an L.D. (lethal dose)-90 of LPS.One series of mice additionally receives phosphate buffered saline (PBS)and Tween 0.5% i.p. 30 minutes before LPS administration. A secondseries consists of groups of mice receiving different doses of the CCR6antagonist given either intra-peritoneally, intra-venously,sub-cutaneously, intra-muscularly, orally, or via any other mode ofadministration 30 minutes before, or concurrently with, LPSadministration. A third series of mice, serving as positive control,consists of groups treated with either mouse IL-10 i.p., or anti-TNFantibodies i.p., 30 minutes before LPS administration. Mice aremonitored for death for 72 hours following the LPS injection.

(e) Evaluation of a Compound of Interest in a Rodent Model ofAsthma/Allergic Lung Inflammation.

CCR6 compounds of the invention can be assessed in a murine model ofallergic asthma. Asthma is induced in 8-10 week old BALB/c mice bysensitizing mice with OVA in Alum adjuvant on days 0 and 10. On day 20mice are challenged with OVA in PBS intranasally to elicit airwayinflammation. Groups of mice are either treated with vehicle, orincreasing doses of a compound of the invention starting on day 20 andlasting until day 23. Animals are subsequently analyzed at day 23 afterthe intranasal OVA challenge for cellular infiltrates in bronchoalveolarlavage (BAL). Mice treated with a compound of the invention will displaysignificantly reduced BAL leukocyte numbers relative to vehicle treatedmice.

(f) Evaluation of a Compound of Interest in a Mouse Model of RheumatoidArthritis.

This example describes a procedure to evaluate the efficacy of CCR6antagonists for treatment of rheumatoid arthritis. An animal model ofrheumatoid arthritis was induced in rodents by injecting them with typeII collagen in selected adjuvants. Three series of rodent groupsconsisting of 15 genetically-susceptible mice or rats per group wereinjected sub-cutaneously or intra-dermally with type II collagenemulsified in Complete Freund's Adjuvant at days 0 and 21. One series ofrodents additionally received PBS and Tween 0.5% i.p. at the initialsensitization and at different dosing schedules thereafter. A secondseries consists of groups of rodents received different doses of theCCR6 antagonist given either intra-peritoneally, intra-venously,sub-cutaneously, intramuscularly, orally, or via any other mode ofadministration at the initial sensitization, and at different dosingschedules thereafter. A third series of rodents, serving as positivecontrol, may consist of groups treated with either mouse IL-10 i.p., oranti-TNF antibodies i.p. at the initial sensitization, and at differentdosing schedules thereafter. Animals were monitored from weeks 3 till 8for the development of swollen joints or paws, and graded on a standarddisease severity scale. Disease severity was confirmed by histologicalanalysis of joints. Animals treated with compound 1.061 of the inventiondisplayed significantly improved scores following treatment.

(g) Evaluation of a Compound of Interest in a Mouse Model of SLE.

This example describes a procedure to evaluate efficacy of CCR6antagonists for treatment of Systemic Lupus Erythematosus (SLE). FemaleNZB/W FI mice spontaneously develop an SLE-like pathology commencing at6 months of age that is characterized by proteinuria, serumautoantibodies, glomerulonephritis, and eventually death. Three seriesof NZB/W FI mouse groups comprising 20 mice per group are tested forefficacy of CCR6 antagonist as follows: One series of mice additionallyreceives phosphate buffered saline (PBS) and Tween 0.5% i.p. soon afterweaning, and thereafter at varying dosing schedules. A second seriesconsists of groups of mice receiving different doses of the CCR6antagonist given either intra-peritoneally, intra-venously,sub-cutaneously, intramuscularly, orally, or via any other mode ofadministration soon after weaning, and thereafter at varying dosingschedules. A third series of mice, serving as positive control, consistsof groups treated with anti-IL10 antibodies given soon after weaning,and thereafter at varying dosing schedules. Disease development ismonitored in terms of eventual mortality, kidney histology, serumautoantibody levels, and proteinuria.

(h) Evaluation of a Compound of Interest in a Mouse Model of Malignancy.

This example describes a procedure to evaluate efficacy of CCR6antagonists for treatment of malignancy. Normal mouse strains can betransplanted with a variety of well-characterized mouse tumor lines,including a mouse thymoma EL4 which has been transfected with OVA toallow easy evaluation of tumor specific antigen responses followingvaccination with OVA. Three series of mouse groups from any of thesetumor models are tested for CCR6 antagonist efficacy as follows: Oneseries of mice additionally receives PBS and Tween 0.5% i.p. soon aftertumor transplant, and thereafter at varying dosing schedules. A secondseries consists of groups of mice receiving different doses of the CCR6antagonist given either intra-peritoneally, intra-venously,sub-cutaneously, intramuscularly, orally, or via any other mode ofadministration soon after tumor transplant, and thereafter at varyingdosing schedules. A third series of mice, serving as positive control,consists of groups treated with either anti-IL17 antibodies, anti-IFNgantibodies, given i.p. soon after tumor transplant, and thereafter atvarying dosing schedules. Efficacy is monitored via tumor growth versusregression. In the case of the OVA-transfected EL4 thymoma model,cytolytic OVA-specific responses can be measured by stimulating draininglymph node cells with OVA in vitro, and measuring antigen-specificcytotoxicity at 72 hours. Analysis can be performed on eg. tumor mass,IL-17 levels or infiltration of regulatory T cells and treatment with acompound of the invention would be expected to produce a beneficialeffect in this model.

(i) Evaluation of a Compound of Interest in a Mouse Carcinoma Model.

The mouse RENCA tumor model accurately mimics the progression of humanadult renal cell carcinoma specifically with reference to spontaneousmetastasis to lungs and serves as a model for solid tumors. Balb/c 6-8week old female mice are inoculated with approximately 5e5 RENCA cells(mouse renal adenocarcinoma; ATCC cat# CRL-2947) under the kidneycapsule and kidney tumor growth is observed over 22 days, with lungmetastasis observed as early as day 15. Animals are dosed with eithervehicle or a compound of the invention eg daily subcutaneously, from thetime of tumor implantation to monitor effects on primary growth, or at alater time (eg day 7) to monitor the compound effect on metastasis.Primary tumor areas are measured twice a week using mechanical calipers.Tumor volumes are calculated by the formula v=pab2/6, where a is thelongest diameter and b is the next longest diameter perpendicular to a.A reduction in tumor volume or incidence of metastasis indicatesefficacy of compound in this indication.

(j) Evaluation of a Compound of Interest in a Mouse Model of IBD.

This example describes a procedure to evaluate the efficacy of CCR6antagonists in Inflammatory Bowel Disease (IBD). Several mouse models ofIBD (including Crohn's Disease and Ulcerative Colitis) have beendeveloped. Some of these are spontaneous models occurring in geneticallyengineered transgenic mice that have been depleted of certain cytokinegenes (e.g. IL-I0, or IL-2). Another mouse model of IBD is obtained bytransferring highly purified populations of CD4+ T lymphocytes bearing aparticular surface marker phenotype (namely CD45 RB hi) into SCID mice.Three series of mouse groups from anyone of these models can be used toevaluate CCR6 antagonist efficacy as follows. One group of miceadditionally receives PBS and Tween 0.5% i.p. soon after weaning in thecase of the spontaneous models in transgenic mice, or at time of celltransfer into SCID mice and varying dosings thereafter for the celltransfer model. A second series consists of groups of mice receivingdifferent doses of the CCR6 antagonist given either intraperitoneally,intra-venously, sub-cutaneously, intra-muscularly, orally, or via anyother mode of administration soon after weaning in the case of thespontaneous models in transgenic mice, or at time of cell transfer intoSCID mice and varying dosings thereafter for the cell transfer model. Athird series of mice, serving as positive control, consists of groupstreated with antibodies to either IFNg, or TNF, or with cytokine IL-10soon after weaning in the case of the spontaneous models in transgenicmice, or at time of cell transfer into SCID mice and varying dosingsthereafter for the cell transfer model. Mice are evaluated for 6-8 weeksfor disease development, monitored initially via weight loss and/orprolapsed rectum, and eventually by histological evaluation of theanimals colon and intestinal tract.

Human CCR6 mRNA transcript variant 1. ACCESSION NM_004367VERSION NM_004367.5 GI:150417991 SEQ ID NO: 1    1agtgtatggg tgaaggaggc agcagtgtgg ccggagagga gagctgggct gggagcacag   61gaaggtcccc aggactctgt ggtcatcagt aagagagggc ccacgtgtat atgctggtga  121acagaaatgt caaccttttc aaagtctgac atttaagaga aaaaactgtg gctgttggtt  181tgtggaacag acagctcctt ctttattgag tcacctctac tttcctgcta ccgctgcctg  241tgagctgaag gggctgaacc atacactcct ttttctacaa ccagcttgca ttttttctgc  301ccacaatgag cggggaatca atgaatttca gcgatgtttt cgactccagt gaagattatt  361ttgtgtcagt caatacttca tattactcag ttgattctga gatgttactg tgctccttgc  421aggaggtcag gcagttctcc aggctatttg taccgattgc ctactccttg atctgtgtct  481ttggcctcct ggggaatatt ctggtggtga tcacctttgc tttttataag aaggccaggt  541ctatgacaga cgtctatctc ttgaacatgg ccattgcaga catcctcttt gttcttactc  601tcccattctg ggcagtgagt catgccaccg gtgcgtgggt tttcagcaat gccacgtgca  661agttgctaaa aggcatctat gccatcaact ttaactgcgg gatgctgctc ctgacttgca  721ttagcatgga ccggtacatc gccattgtac aggcgactaa gtcattccgg ctccgatcca  781gaacactacc gcgcagcaaa atcatctgcc ttgttgtgtg ggggctgtca gtcatcatct  841ccagctcaac ttttgtcttc aaccaaaaat acaacaccca aggcagcgat gtctgtgaac  901ccaagtacca gactgtctcg gagcccatca ggtggaagct gctgatgttg gggcttgagc  961tactctttgg tttctttatc cctttgatgt tcatgatatt ttgttacacg ttcattgtca 1021aaaccttggt gcaagctcag aattctaaaa ggcacaaagc catccgtgta atcatagctg 1081tggtgcttgt gtttctggct tgtcagattc ctcataacat ggtcctgctt gtgacggctg 1141caaatttggg taaaatgaac cgatcctgcc agagcgaaaa gctaattggc tatacgaaaa 1201ctgtcacaga agtcctggct ttcctgcact gctgcctgaa ccctgtgctc tacgctttta 1261ttgggcagaa gttcagaaac tactttctga agatcttgaa ggacctgtgg tgtgtgagaa 1321ggaagtacaa gtcctcaggc ttctcctgtg ccgggaggta ctcagaaaac atttctcggc 1381agaccagtga gaccgcagat aacgacaatg cgtcgtcctt cactatgtga tagaaagctg 1441agtctcccta aggcatgtgt gaaacatact catagatgtt atgcaaaaaa aagtctatgg 1501ccaggtatgc atggaaaatg tgggaattaa gcaaaatcaa gcaagcctct ctcctgcggg 1561acttaacgtg ctcatgggct gtgtgatctc ttcagggtgg ggtggtctct gataggtagc 1621attttccagc actttgcaag gaatgttttg tagctctagg gtatatatcc gcctggcatt 1681tcacaaaaca gcctttggga aatgctgaat taaagtgaat tgttgacaaa tgtaaacatt 1741ttcagaaata ttcatgaagc ggtcacagat cacagtgtct tttggttaca gcacaaaatg 1801atggcagtgg tttgaaaaac taaaacagaa aaaaaaatgg aagccaacac atcactcatt 1861ttaggcaaat gtttaaacat ttttatctat cagaatgttt attgttgctg gttataagca 1921gcaggattgg ccggctagtg tttcctctca tttccatttg atacagtcaa caagcctgac 1981cctgtaaaat ggaggtggaa agacaagctc aagtgttcac aacctggaag tgcttcggga 2041agaaggggac aatggcagaa caggtgttgg tgacaattgt caccaattgg ataaagcagc 2101tcaggttgta gtgggccatt aggaaactgt cggtttgctt tgatttccct gggagctgtt 2161ctctgtcgtg agtgtctctt gtctaaacgt ccattaagct gagagtgcta tgaagacagg 2221atctagaata atcttgctca cagctgtgct ctgagtgcct agcggagttc cagcaaacaa 2281aatggactca agagagattt cattaatgaa tcgtaatgaa gttggggttt attgtacagt 2341ttaaaatgtt agatgttttt aattttttaa ataaatggaa tacttttttt ttttttttaa 2401agaaagcaac tttactgaga caatgtagaa agaagttttg ttccgtttct ttaatgtggt 2461tgaagagcaa tgtgtggctg aagacttttg ttatgaggag ctgcagatta gctaggggac 2521agctggaatt atgctggctt ctgataatta ttttaaaggg gtctgaaatt tgtgatggaa 2581tcagatttta acagctctct tcaatgacat agaaagttca tggaactcat gtttttaaag 2641ggctatgtaa atatatgaac attagaaaaa tagcaacttg tgttacaaaa atacaaacac 2701atgttaggaa ggtactgtca tgggctaggc atggtggctc acacctgtaa tcccagcatt 2761ttgggaagct aagatgggtg gatcacttga ggtcaggagt ttgagaccag cctggccaac 2821atggcgaaac ccctctctac taaaaataca aaaatttgcc aggcgtggtg gcgggtgcct 2881gtaatcccag ctacttggga ggctgaggca agagaatcgc ttgaacccag gaggcagagg 2941ttgcagtgag ccgagatcgt gccattgcac tccagcctgg gtgacaaagc gagactccat 3001ctcaaaaaaa aaaaaaaaaa aaaaggaaag aactgtcatg taaacatacc aacatgttta 3061aacctgacaa tggtgttatt tgaaacttta tattgttctt gtaagcttta actatatctc 3121tctttaaaat gcaaaataat gtcttaagat tcaaagtctg tatttttaaa gcatggcttt 3181ggctttgcaa aataaaaaat gtgttttgta catgaaHuman CCR6 mRNA transcript variant 2. ACCESSION NM_031409VERSION NM_031409.3 GI:150417990 SEQ ID NO: 2    1aactcacacg gcctcttgca aacgttccca aatcttccca gtcggcttgc agagactcct   61tgctcccagg agataaccag gtaaaggagt atgaaagttt gggtacaaac tcattgctgc  121aaattgaaaa ccatgcaaag gctgtcttcc tctggggagt tcaatgcctc tcttttactt  181atcactttac cattggttgg actttgattc cagggatcct acgattactc aataccctac  241aggatataca tggttaacca tttgcatttg ggcaaatagg cgttactttt caataggaag  301tggcaatcca gaacttgctt ttgggcaatt ctagtagctc accgcttttt tcttaatgac  361tgctagaagc tgcatcttat tgacagatgg tcatcacatt ggtgagctgg agtcatcaga  421ttgtggggcc cggagtgagg ctgaagggag tggatcagag cactgcctga gagtcacctc  481tactttcctg ctaccgctgc ctgtgagctg aaggggctga accatacact catttttcta  541caaccagctt gcattttttc tgcccacaat gagcggggaa tcaatgaatt tcagcgatgt  601tttcgactcc agtgaagatt attttgtgtc agtcaatact tcatattact cagttgattc  661tgagatgtta ctgtgctcct tgcaggaggt caggcagttc tccaggctat ttgtaccgat  721tgcctactcc ttgatctgtg tctttggcct cctggggaat attctggtgg tgatcacctt  781tgctttttat aagaaggcca ggtctatgac agacgtctat ctcttgaaca tggccattgc  841agacatcctc tttgttctta ctctcccatt ctgggcagtg agtcatgcca ccggtgcgtg  901ggttttcagc aatgccacgt gcaagttgct aaaaggcatc tatgccatca actttaactg  961cgggatgctg ctcctgactt acattaacat ggaccggtac atcgccattg tacaggcgac 1021taagtcattc cggctccgat ccagaacact accgcgcagc aaaatcatct gccttgttgt 1081gtgggggctg tcagtcatca tctccagctc aacttttgtc ttcaaccaaa aatacaacac 1141ccaaggcagc gatgtctgtg aacccaagta ccagactgtc tcggagccca tcaggtggaa 1201gctgctgatg ttggggcttg agctactctt tggtttcttt atccctttga tgttcatgat 1261attttgttac acgttcattg tcaaaacctt ggtgcaagct cagaattcta aaaggcacaa 1321agccatccgt gtaatcatag ctgtggtgct tgtgtttctg gcttgtcaga ttcctcataa 1381catggtcctg cttgtgacgg ctgcaaattt gggtaaaatg aaccgatcct gccagagcga 1441aaagctaatt ggctatacga aaactgtcac agaagtcctg gctttcctgc actgctgcct 1501gaaccctgtg ctctacgctt ttattgggca gaagttcaga aactactttc tgaagatctt 1561gaaggacctg tggtgtgtga gaaggaagta caagtcctca ggcttctcct gtgccgggag 1621gtactcagaa aacatttctc ggctgaccag tgagaccgca gataacgaca atgcgtcgtc 1681cttcactatg tgatagaaag ctgagtctcc ctaaggcatg tgtgaaacat actcatagat 1741gttatgcaaa aaaaagtcta tggccaggta tgcatggaaa atgtgggaat taagcaaaat 1801caagcaagcc tctctcctgc gggacttaac gtgctcatgg gctgtgtgat ctcttcaggg 1861tggggtggtc tctgataggt agcattttcc agcactttgc aaggaatgtt ttgtagctct 1921agggtatata tccgcctggc atttcacaaa acagcctttg ggaaatgctg aattaaagtg 1981aattgttgac aaatgtaaac attttcagaa atattcatga agcggtcaca gatcacagtg 2041tcttttggtt acagcacaaa atgatggcag tggtttgaaa aactaaaaca gaaaaaaaaa 2101tggaagccaa cacatcactc attttaggca aatgtttaaa catttttatc tatcagaatg 2161tttattgttg ctggttataa gcagcaggat tggccggcta gtgtttcctc tcatttccct 2221ttgatacagt caacaagcct gaccctgtaa aatggaggtg gaaagacaag ctcaagtgtt 2281cacaacctgg aagtgcttcg ggaagaaggg gacaatggca gaacaggtgt tggtgacaat 2341tgtcaccaat tggataaagc agctcaggtt gtagtgggcc attaggaaac tgtcggtttg 2401ctttgatttc cctgggagct gttctctgtc gtgagtgtct cttgtctaaa cgtccattaa 2461gctgagagtg ctatgaagac aggatctaga ataatcttgc tcacagctgt gctctgagtg 2521cctagcggag ttccagcaaa caaaatggac tcaagagaga tttgattaat gaatcgtaat 2581gaagttgggg tttattgtac agtttaaaat gttagatgtt tttaattttt taaataaatg 2641gaatactttt tttttttttt taaagaaagc aactttactg agacaatgta gaaagaagtt 2701ttgttccgtt tctttaatgt ggttgaagag caatgtgtgg ctgaagactt ttgttatgag 2761gagctgcaga ttagctaggg gacagctgga attatgctgg cttctgataa ttattttaaa 2821ggggtctgaa atttgtgatg gaatcagatt ttaacagctc tcttcaatga catagaaagt 2881tcatggaact catgttttta aagggctatg taaatatatg aacattagaa aaatagcaac 2941ttgtgttaca aaaatacaaa cacatgttag gaaggtactg tcatgggcta ggcatggtgg 3001ctcacacctg taatcccagc attttgggaa gctaagatgg gtggatcact tgaggtcagg 3061agtttgagac cagcctggcc aacatggcga aacccctctc tactaaaaat acaaaaattt 3121gccaggcgtg gtggcgggtg cctgtaatcc cagctacttg ggaggctgag gcaagagaat 3181cgcttgaacc caggaggcag aggttgcagt gagccgagat cgtgccattg cactccagcc 3241tgggtgacaa agcgagactc catcttaaaa aaaaaaaaaa aaaaaaagga aagaactgtc 3301atgtaaacat accaacatgt ttaaacctga caatggtgtt atttgaaact ttatattgtt 3361cttgtaagct ttaactatat ctctctttaa aatgcaaaat aatgtcttaa gattcaaagt 3421ctgtattttt aaagcatggc tttggctttg caaaataaaa aatgtgtttt gtacatgaaHuman CCR6 amino acid sequence SEQ ID NO: 3MSGESMNFSDVFDSSEDYFVSVNTSYYSVDSEMLLCSLQEVRQFSRLFVPIAYSLICVFGLLGNILVVITFAFYKKARSMTDVYLLNMAIADILFVLTLPFWAVSHATGAWVFSNATCKLLKGIYAINFNCGMLLLTCISMDRYIAIVQATKSFRLRSRTLPRSKIICLVVWGLSVIISSSTFVFNQKYNTQGSDVCEPKYQTVSEPIRWKLLMLGLELLFGFFIPLMFMIFCYTFIVKTLVQAQNSKRHKAIRVIIAVVLVFLACQIPHNMVLLVTAANLGKMNRSCQSEKLIGYTKTVTEVLAFLHCCLNPVLYAFIGQKRNYFLKILKDLWCVRRKYKSSGFSCAGRYSENISRQTSETADNDNASSFTM

It is understood that the examples and embodiments described herein arefor illustrative purposes only and that various modifications or changesin light thereof will be suggested to persons skilled in the art and areto be included within the spirit and purview of this application andscope of the appended claims.

All publications, patents, and patent applications cited herein arehereby incorporated by reference in their entirety for all purposes.

What is claimed is:
 1. A compound having the formula

or a pharmaceutically acceptable salt thereof.
 2. A compound of claim 1,having the formula


3. A compound having the formula

or a pharmaceutically acceptable salt thereof.
 4. A compound of claim 3,having the formula


5. A compound having the formula

or a pharmaceutically acceptable salt thereof.
 6. A compound of claim 5,having the formula